Articoli di riviste: "Substrate identification"

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Malintoi, Adrianus, Inneke F. M. Rumengan, Kakaskasen A. Roeroe, Veibe Warouw, Ari B. Rondonuwu e Medy Ompi. "KOMUNITAS ASCIDIA DI PESISIR MALALAYANG DUA, TELUK MANADO, SULAWESI UTARA". JURNAL PESISIR DAN LAUT TROPIS 8, n. 1 (15 gennaio 2020): 39. http://dx.doi.org/10.35800/jplt.8.1.2020.27403.

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Field survey on ascidian community was conducted along the coastal area of Malalayang Dua in order to find out species of ascidia, species abundance, and ascidian substrates. A survey method and quadrant transects were applied. Pictures were taken, while species and their substrates were sampled. Species identification was based on morphological characteristics, while substrate type identification was based on ascidian species attachment. The results shows that 21 ascidian species were found in the the coastal of Malalayang Dua. Didemnum molle was the highest abundant species in the area, followed by Polycarpa aurata, Polycarpa sp.4. and Polycarpa sp.2.. Dead coral algaes (DCA) were found to be the most preferred substrates by ascidians in the area. Keywords : ascidia, species, substrate, distribution, and abundance Survei lapangan terhadap komunitas ascidia dilakukan di pesisir Malalayang Dua untuk mendapatkan data jenis, kelimpahan, dan substrat ascidia. Metode yang digunakan yaitu metode survei jelajah dan transek kuadran. Identifikasi jenis ascidia dilakukan berdasarkan karakteristik morfologi. Hasil penelitian ditemukan ada 21 jenis ascidia. Substrat jenis death coral algae (DCA) merupakan substrat yang paling banyak ditempati ascidia. Kelimpahan ascidia tertinggi adalah Didemnum molle di pesisir Malalayang Dua, diikuti oleh Polycarpa aurata, Polycarpa sp.4. dan Polycarpa sp.2. Death coral alga (DCA) ditemukan sebagai substrat yang paling disukai oleh ascidia di daerah itu. Kata Kunci : ascidia, spesies, substrat, distribusi, dan kelimpahan

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Baros, Seanantha S., Jonathan M. Blackburn e Nelson C. Soares. "Phosphoproteomic Approaches to Discover Novel Substrates of Mycobacterial Ser/Thr Protein Kinases". Molecular & Cellular Proteomics 19, n. 2 (15 dicembre 2019): 233–44. http://dx.doi.org/10.1074/mcp.r119.001668.

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Mycobacterial Ser/Thr protein kinases (STPKs) play a critical role in signal transduction pathways that ultimately determine mycobacterial growth and metabolic adaptation. Identification of key physiological substrates of these protein kinases is, therefore, crucial to better understand how Ser/Thr phosphorylation contributes to mycobacterial environmental adaptation, including response to stress, cell division, and host-pathogen interactions. Various substrate detection methods have been employed with limited success, with direct targets of STPKs remaining elusive. Recently developed mass spectrometry (MS)-based phosphoproteomic approaches have expanded the list of potential STPK substrate identifications, yet further investigation is required to define the most functionally significant phosphosites and their physiological importance. Prior to the application of MS workflows, for instance, GarA was the only known and validated physiological substrate for protein kinase G (PknG) from pathogenic mycobacteria. A subsequent list of at least 28 candidate PknG substrates has since been reported with the use of MS-based analyses. Herein, we integrate and critically review MS-generated datasets available on novel STPK substrates and report new functional and subcellular localization enrichment analyses on novel candidate protein kinase A (PknA), protein kinase B (PknB) and PknG substrates to deduce the possible physiological roles of these kinases. In addition, we assess substrate specificity patterns across different mycobacterial STPKs by analyzing reported sets of phosphopeptides, in order to determine whether novel motifs or consensus regions exist for mycobacterial Ser/Thr phosphorylation sites. This review focuses on MS-based techniques employed for STPK substrate identification in mycobacteria, while highlighting the advantages and challenges of the various applications.

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Peerce, B. E. "Identification of the intestinal Na-phosphate cotransporter". American Journal of Physiology-Gastrointestinal and Liver Physiology 256, n. 4 (1 aprile 1989): G645—G652. http://dx.doi.org/10.1152/ajpgi.1989.256.4.g645.

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Na-dependent phosphate uptake in intestinal brush-border membrane vesicles was sensitive to arginine group-specific reagents in a substrate-sensitive manner. Four different arginine group-specific reagents were tested. All four reagents irreversibly inhibited Na-dependent phosphate uptake with the concentration for 50% inhibition, K0.5, varying between 150 and 40 microM. Maximum inhibition approached 80%. Addition of substrates during exposure to these reagents resulted in protection of Na-phosphate cotransport only in the presence of Na and phosphate. Na-phosphate cotransporter labeling at or near the phosphate site was accomplished using a pretreatment step with phenylglyoxal and substrates followed by a fluorescent phenylglyoxal-labeling step. Fluorescein isothiocyanate-phenylglyoxal (FITC-PG) specifically labeled a 130-kDa polypeptide on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) in a substrate-sensitive manner, consistent with its effect on Na-phosphate cotransport. n-Acetylimidazole (NAI) inhibited Na-phosphate cotransport in a Na+- but not K+ -sensitive manner. NAI or fluorescein n-acetylimidazole (FNAI) inhibited Na-dependent phosphate uptake with a K0.5 for inhibition of 38 microM. Maximum inhibition of Na-phosphate cotransport was 75%. On SDS-PAGE, FNAI labeled five polypeptide bands in a Na-sensitive manner including the 130-kDa polypeptide band labeled by FITC-PG. Of these five bands only the 130-kDa polypeptide lost substrate protectability against FITC-PG inhibition of Na-phosphate cotransport and FITC-PG labeling on prior exposure to NAI in the absence of Na+. On this basis the 130-kDa polypeptide is tentatively identified as the intestinal Na-phosphate cotransporter and this polypeptide band contains both the Na substrate site and the phosphate substrate site.

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Gopalakrishnan, Ramakrishnan, Sivakumar Rajagopal, Sai Viswanth Reddy e Anirudh E. R. "Identification of Most Suitable Dielectrics Substrate for UWB Bandpass Filter". ECS Transactions 107, n. 1 (24 aprile 2022): 431–38. http://dx.doi.org/10.1149/10701.0431ecst.

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Identification of a suitable substrate is a critical parameter in the design of microstrip bandpass filters. In this work, the performance of an Ultra Wideband (UWB) bandpass filter based on Electromagnetic Bandgap (EBG) resonators are designed, fabricated, and analyzed. Printed Circuit Board (PCB) with suitable dielectric substrates are used extensively to develop microstrip lines. The filter is fabricated using RT Duroid substrate material and compared with FR4 and Alumina dielectric substrates and the performance results show that the best result is obtained with RT Duroid.

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SONG, JIANGNING, HAO TAN, SARAH E. BOYD, HONGBIN SHEN, KHALID MAHMOOD, GEOFFREY I. WEBB, TATSUYA AKUTSU, JAMES C. WHISSTOCK e ROBERT N. PIKE. "BIOINFORMATIC APPROACHES FOR PREDICTING SUBSTRATES OF PROTEASES". Journal of Bioinformatics and Computational Biology 09, n. 01 (febbraio 2011): 149–78. http://dx.doi.org/10.1142/s0219720011005288.

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Proteases have central roles in "life and death" processes due to their important ability to catalytically hydrolyze protein substrates, usually altering the function and/or activity of the target in the process. Knowledge of the substrate specificity of a protease should, in theory, dramatically improve the ability to predict target protein substrates. However, experimental identification and characterization of protease substrates is often difficult and time-consuming. Thus solving the "substrate identification" problem is fundamental to both understanding protease biology and the development of therapeutics that target specific protease-regulated pathways. In this context, bioinformatic prediction of protease substrates may provide useful and experimentally testable information about novel potential cleavage sites in candidate substrates. In this article, we provide an overview of recent advances in developing bioinformatic approaches for predicting protease substrate cleavage sites and identifying novel putative substrates. We discuss the advantages and drawbacks of the current methods and detail how more accurate models can be built by deriving multiple sequence and structural features of substrates. We also provide some suggestions about how future studies might further improve the accuracy of protease substrate specificity prediction.

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Dauksher, Walter, Scott Burton, David Niles e Dennis H. Eaton. "Identification of Poor Via-Ceramic Adhesion in Electronic Substrates". EDFA Technical Articles 4, n. 1 (1 febbraio 2002): 5–10. http://dx.doi.org/10.31399/asm.edfa.2002-1.p005.

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Abstract Mechanical bending strength measurements, part of a quality screening process for ceramic substrate package designs, revealed a significant difference between two substrates that could not be attributed to constituent materials or structural thickness effects. Structural analysis was performed to generate a hypothesis that the metal vias were not structurally adhered to the ceramic holes and thus caused stress concentrations that reduced the bending strength of the substrates. In this detailed case study, the authors explain how they proved their hypothesis using FEA to calculate stress concentration factors, optical microscopy to examine fracture surfaces, and electron spectroscopy to determine chemical compositions. The successful outcome attests to the value of strength criteria as an investigative tool for the assessment of electronic package substrate quality.

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Mizunuma, Masataka, Atsushi Kaneko, Shunta Imai, Kazuhiro Furukawa e Yoshiro Chuman. "Methods for Identification of Substrates/Inhibitors of FCP/SCP Type Protein Ser/Thr Phosphatases". Processes 8, n. 12 (4 dicembre 2020): 1598. http://dx.doi.org/10.3390/pr8121598.

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Protein phosphorylation is the most widespread type of post-translational modification and is properly controlled by protein kinases and phosphatases. Regarding the phosphorylation of serine (Ser) and threonine (Thr) residues, relatively few protein Ser/Thr phosphatases control the specific dephosphorylation of numerous substrates, in contrast with Ser/Thr kinases. Recently, protein Ser/Thr phosphatases were reported to have rigid substrate recognition and exert various biological functions. Therefore, identification of targeted proteins by individual protein Ser/Thr phosphatases is crucial to clarify their own biological functions. However, to date, information on the development of methods for identification of the substrates of protein Ser/Thr phosphatases remains scarce. In turn, substrate-trapping mutants are powerful tools to search the individual substrates of protein tyrosine (Tyr) phosphatases. This review focuses on the development of novel methods for the identification of Ser/Thr phosphatases, especially small C-terminal domain phosphatase 1 (Scp1), using peptide-displayed phage library with AlF4−/BeF3−, and discusses the identification of putative inhibitors.

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Ye, Siying, Siew Yeen Chai, Rebecca A. Lew, David B. Ascher, Craig J. Morton, Michael W. Parker e Anthony L. Albiston. "Identification of modulating residues defining the catalytic cleft of insulin-regulated aminopeptidase". Biochemistry and Cell Biology 86, n. 3 (aprile 2008): 251–61. http://dx.doi.org/10.1139/o08-037.

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Inhibition of insulin-regulated aminopeptidase (IRAP) has been demonstrated to facilitate memory in rodents, making IRAP a potential target for the development of cognitive enhancing therapies. In this study, we generated a 3-D model of the catalytic domain of IRAP based on the crystal structure of leukotriene A4 hydrolase (LTA4H). This model identified two key residues at the ‘entrance’ of the catalytic cleft of IRAP, Ala427 and Leu483, which present a more open arrangement of the S1 subsite compared with LTA4H. These residues may define the size and 3-D structure of the catalytic pocket, thereby conferring substrate and inhibitor specificity. Alteration of the S1 subsite by the mutation A427Y in IRAP markedly increased the rate of substrate cleavage V of the enzyme for a synthetic substrate, although a corresponding increase in the rate of cleavage of peptide substrates Leu-enkephalin and vasopressin was was not apparent. In contrast, [L483F]IRAP demonstrated a 30-fold decrease in activity due to changes in both substrate affinity and rate of substrate cleavage. [L483F]IRAP, although capable of efficiently cleaving the N-terminal cysteine from vasopressin, was unable to cleave the tyrosine residue from either Leu-enkephalin or Cyt6-desCys1-vasopressin (2–9), both substrates of IRAP. An 11-fold reduction in the affinity of the peptide inhibitor norleucine1-angiotensin IV was observed, whereas the affinity of angiotensin IV remained unaltered. In additionm we predict that the peptide inhibitors bind to the catalytic site, with the NH2-terminal P1 residue occupying the catalytic cleft (S1 subsite) in a manner similar to that proposed for peptide substrates.

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Zhao, Yi, Eliud Morales Morales Dávila, Xue Li, Beiyu Tang, Ariana I. Rabinowitsch, Jose Manuel Perez-Aguilar e Carl P. Blobel. "Identification of Molecular Determinants in iRhoms1 and 2 That Contribute to the Substrate Selectivity of Stimulated ADAM17". International Journal of Molecular Sciences 23, n. 21 (24 ottobre 2022): 12796. http://dx.doi.org/10.3390/ijms232112796.

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The metalloprotease ADAM17 is a key regulator of the TNFα, IL-6R and EGFR signaling pathways. The maturation and function of ADAM17 is controlled by the seven-membrane-spanning proteins iRhoms1 and 2. The functional properties of the ADAM17/iRhom1 and ADAM17/iRhom2 complexes differ, in that stimulated shedding of most ADAM17 substrates tested to date can be supported by iRhom2, whereas iRhom1 can only support stimulated shedding of very few ADAM17 substrates, such as TGFα. The first transmembrane domain (TMD1) of iRhom2 and the sole TMD of ADAM17 are important for the stimulated shedding of ADAM17 substrates by iRhom2. However, little is currently known about how the iRhoms interact with different substrates to control their stimulated shedding by ADAM17. To provide new insights into this topic, we tested how various chimeras between iRhom1 and iRhom2 affect the stimulated processing of the EGFR-ligands TGFα (iRhom1- or 2-dependent) and EREG (iRhom2-selective) by ADAM17. This uncovered an important role for the TMD7 of the iRhoms in determining their substrate selectivity. Computational methods utilized to characterize the iRhom1/2/substrate interactions suggest that the substrate selectivity is determined, at least in part, by a distinct accessibility of the substrate cleavage site to stimulated ADAM17. These studies not only provide new insights into why the substrate selectivity of stimulated iRhom2/ADAM17 differs from that of iRhom1/ADAM17, but also suggest new approaches for targeting the release of specific ADAM17 substrates.

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Zhai, Jingyu, Yugang Chen, Xinyuan Song, Hongchun Wu e Qingkai Han. "Identification of the Anisotropic Elastic Parameters of NiCrAlY Coating by Combining Nanoindentation and Finite Element Method". Shock and Vibration 2019 (2 giugno 2019): 1–13. http://dx.doi.org/10.1155/2019/9034750.

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For vibration damping, coatings are prepared on surface of the structures (substrates), which constitute the coating-substrate composite structures. Elastic parameters of the coating are indispensable for the vibration and damping analysis of the composite structure. Due to the small scale of coating thickness and elastic difference compared with the substrate, the identification results are inevitably influenced by the existence of substrate. Moreover, resulting from the preparation process, elastic properties of hard coating often exhibit anisotropic properties. All the above factors bring about the difficulties of accurate identification. In this study, a method for identifying anisotropic elastic parameters of hard coatings considering substrate effect is proposed, by combining nanoindentation and finite element analysis. Based on the identification results, finite element models are established to analyze the vibration characteristics of the coating-substrate composite structure, which verify the rationality of the anisotropic elastic parameters for vibration analysis. The studies in this paper are significant to more accurately identify the mechanical parameters for establishing the dynamic model. Moreover, they lay the foundation for further optimization design of hard coating damping.

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Garton, A. J., A. J. Flint e N. K. Tonks. "Identification of p130(cas) as a substrate for the cytosolic protein tyrosine phosphatase PTP-PEST." Molecular and Cellular Biology 16, n. 11 (novembre 1996): 6408–18. http://dx.doi.org/10.1128/mcb.16.11.6408.

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PTP-PEST is a ubiquitously expressed, cytosolic, mammalian protein tyrosine phosphatase (PTP) which exhibits high specific activity in vitro. We have investigated the substrate specificity of PTP-PEST by a novel substrate-trapping approach in combination with in vitro dephosphorylation experiments. We initially identified a prominent 130-kDa tyrosine-phosphorylated protein in pervanadate-treated HeLa cell lysates which was preferentially dephosphorylated by PTP-PEST in vitro. In order to identify this potential substrate, mutant (substrate-trapping) forms of PTP-PEST were generated which lack catalytic activity but retain the ability to bind substrates. These mutant proteins associated in stable complexes exclusively with the same 130-kDa protein, which was identified as p130(cas) by immunoblotting. This exclusive association was observed in lysates from several cell lines and in transfected COS cells, but was not observed with other members of the PTP family, strongly suggesting that p130(cas) represents a major physiologically relevant substrate for PTP-PEST. Our studies suggest potential roles for PTP-PEST in regulation of p130(cas) function. These functions include mitogen- and cell adhesion-induced signalling events and probable roles in transformation by various oncogenes. These results provide the first demonstration of a PTP having an inherently restricted substrate specificity in vitro and in vivo. The methods used to identify p130(cas) as a specific substrate for PTP-PEST are potentially applicable to any PTP and should therefore prove useful in determining the physiological substrates of other members of the PTP family.

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Peng, C., A. Knebel, N. A. Morrice, X. Li, K. Barringer, J. Li, S. Jakes, B. Werneburg e L. Wang. "Pim Kinase Substrate Identification and Specificity". Journal of Biochemistry 141, n. 3 (19 dicembre 2006): 353–62. http://dx.doi.org/10.1093/jb/mvm040.

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Luo, Shu Yue, Luam Ellen Araya e Olivier Julien. "Protease Substrate Identification Using N-terminomics". ACS Chemical Biology 14, n. 11 (agosto 2019): 2361–71. http://dx.doi.org/10.1021/acschembio.9b00398.

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Fujimoto, Tomohito, Naoya Hatano, Naohito Nozaki, Saki Yurimoto, Ryoji Kobayashi e Hiroshi Tokumitsu. "Identification of a novel CaMKK substrate". Biochemical and Biophysical Research Communications 410, n. 1 (giugno 2011): 45–51. http://dx.doi.org/10.1016/j.bbrc.2011.05.102.

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Garnica B., Sergio A., Marius Knaust e Sergej Fatikow. "Automatic Micro-Robotic Identification and Electrical Characterization of Graphene". Micromachines 10, n. 12 (11 dicembre 2019): 870. http://dx.doi.org/10.3390/mi10120870.

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Micromechanically exfoliating graphene on S i / S i O 2 substrates is commonplace for graphene researchers, but locating actual graphene flakes on these substrates is a high-effort and tiresome task. The main purpose of this work was to establish a completely automated procedure to identify those graphene flakes with as little human interaction as possible, improving on the limitations of current methods. Furthermore, automatic electrical characterization of the identified flakes was performed. The proposed micro-robotic automation sequence consists of three main steps. To start, a sample surface plane is calculated, based on multiple foci points across the substrate. Secondly, flakes on the substrate are identified in the hue, saturation, and value (HSV) color space, with an implementation to fit the measurement probe, used to avoid undersized samples and adjust the flake orientation. Finally, electrical characterization is performed based on four point probe measurements with the Van der Pauw method. Results of the successfully implemented automation sequence are presented together with flake electrical properties and validation.

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Renaud, François N. R., Marianne Dutaur, Salah Daoud, Dominique Aubel, Philippe Riegel, Daniel Monget e Jean Freney. "Differentiation of Corynebacterium amycolatum, C. minutissimum, and C. striatum by Carbon Substrate Assimilation Tests". Journal of Clinical Microbiology 36, n. 12 (1998): 3698–702. http://dx.doi.org/10.1128/jcm.36.12.3698-3702.1998.

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We tested the carbon substrate assimilation patterns of 40Corynebacterium amycolatum strains, 19 C. minutissimum strains, 50 C. striatum strains, and 1C. xerosis strain with the Biotype 100 system (bioMérieux, Marcy-l’Étoile, France). Twelve carbon substrates of 99 allowed discrimination among the species tested. Additionally, assimilation of 3 of these 12 carbon substrates (maltose, N -acetyl-d-glucosamine, and phenylacetate) was tested with the API 20 NE identification system (bioMérieux). Since concordant results were observed with the two systems for these three carbon substrates, either identification system can be used as a supplementary tool to achieve phenotypic differential identification ofC. amycolatum, C. minutissimum, and C. striatum in the clinical microbiology laboratory.

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Li, Yuanxin, Ningning Dong, Saifeng Zhang, Kangpeng Wang, Long Zhang e Jun Wang. "Optical identification of layered MoS2via the characteristic matrix method". Nanoscale 8, n. 2 (2016): 1210–15. http://dx.doi.org/10.1039/c5nr06287j.

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The characteristic matrix method is demonstrated to be effective and reliable for the calculation of the optical contrast of 2D materials on various substrates. This work provides a guide for the selection of the illumination wavelength or the substrate when observing the 2D materials by using optical microscopy.

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Liz, Márcia A., Carolina E. Fleming, Ana F. Nunes, Maria R. Almeida, Fernando M. Mar, Youngchool Choe, Charles S. Craik, James C. Powers, Matthew Bogyo e Mónica M. Sousa. "Substrate specificity of transthyretin: identification of natural substrates in the nervous system". Biochemical Journal 419, n. 2 (27 marzo 2009): 467–74. http://dx.doi.org/10.1042/bj20082090.

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Besides functioning as the plasma transporter of retinol and thyroxine, TTR (transthyretin) is a protease, cleaving apoA-I (apolipoprotein A-I) after a phenylalanine residue. In the present study, we further investigated TTR substrate specificity. By using both P-diverse libraries and a library of phosphonate inhibitors, a TTR preference for a lysine residue in P1 was determined, suggesting that TTR might have a dual specificity and that, in addition to apoA-I, other TTR substrates might exist. Previous studies revealed that TTR is involved in the homoeostasis of the nervous system, as it participates in neuropeptide maturation and enhances nerve regeneration. We investigated whether TTR proteolytic activity is involved in these functions. Both wild-type TTR and TTRprot− (proteolytically inactive TTR) had a similar effect in the expression of peptidylglycine α-amidating mono-oxygenase, the rate-limiting enzyme in neuropeptide amidation, excluding the involvement of TTR proteolytic activity in neuropeptide maturation. However, TTR was able to cleave amidated NPY (neuropeptide Y), probably contributing to the increased NPY levels reported in TTR-knockout mice. To assess the involvement of TTR proteolytic activity in axonal regeneration, neurite outgrowth of cells cultivated with wild-type TTR or TTRprot−, was measured. Cells grown with TTRprot− displayed decreased neurite length, thereby suggesting that TTR proteolytic activity is important for its function as a regeneration enhancer. By showing that TTR is able to cleave NPY and that its proteolytic activity affects axonal growth, the present study shows that TTR has natural substrates in the nervous system, establishing further its relevance in neurobiology.

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Kim, Seong-Tae, Dae-Sik Lim, Christine E. Canman e Michael B. Kastan. "Substrate Specificities and Identification of Putative Substrates of ATM Kinase Family Members". Journal of Biological Chemistry 274, n. 53 (31 dicembre 1999): 37538–43. http://dx.doi.org/10.1074/jbc.274.53.37538.

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Saragih, Hans S. R. P., Medy Ompi, Erly Yosef Kaligis, Farnis B. Boneka Boneka, Veibe Warouw e Darus Sa’adah Johanis Paransa. "Attachment Of Macrobenthos Larvae To Organic And Non-Organic Substrates". Jurnal Ilmiah PLATAX 12, n. 1 (26 gennaio 2024): 185–93. http://dx.doi.org/10.35800/jip.v12i1.52205.

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The objectives of this study are 1) to Identify the specimen of macrobenthos attached to the substrate provided in 2 locations. 2) to determine the type of substrate attached by the larvae at both sites. 3) to determine the density of Macrobenthos attached to the substrate provided at 2 sites. A plywood plate had 16 holes with a diameter of 1 cm each that had been randomly filled with organic substrates, namely coconut fibers, palm fibers, shells with 'bysus' threads, and non-organic substrates in the form of plastic ropes. Each substrate has four replicates. The plywood plate with 3 replicates was placed in each station. The plate was removed and taken to the laboratory after 1 month. The identification up to family, genus, and species as well as the density of each species were applied. The results showed that not only the larvae of Septifer bilocularis attached to organic and non-organic substrates, but six species of larvae were also identified. The sizes of new settler macrobenthos from 2 mm to 1 cm attached on organic and non-organic substrates were identified. The density of new settlers species attached to substrates varied from 0.03 – 0.5 individuals/cm2. The new settlers identified 7 species in Tiwoho and 3 species in Malalayang. Keywords: Settlement, Substrate, Tiwoho Coast, Malalayang Coas. Abstrak Tujuan dari penelitian ini adalah 1) Mengidentifikasi jenis-jenis makrobenthos yang menempel pada substrat yang disediakan di 2 lokasi. 2) Mengidentifikasi jenis substrat sebagai tempat penempelan larva macrobenthos di kedua lokasi, dan 3) Menentukan kepadatan jenis Makrobenthos pada substrat yang disediakan di 2 lokasi. Triplek (plate) memiliki 16 lubang dengan ukuran diameter masing-masing 1 cm yang telah diisi secara acak dengan substrat organik yaitu serabut kelapa, serabut ijuk, cangkang ber ‘byssus’, serta substrat non organik berupa tali plastik. Masing-masing substrat ini memiliki 4 ulangan. Selanjutnya, plate, masing-masing dengan 3 ulangan ditempatkan di setiap intertidal, Towoho dan Malalayang. Plate diangkat setelah 1 bulan, yang dibawah ke laboratorium untuk foto dan diidentifikasi baik di tingkat jenis, genus, ataupun family. Hasil penelitian menunjukkan bahwa tidak hanya larva kerang Septifer bilocularis (Linnaeus, 1758), yang menempel pada substrat organik dan non organik, tetapi ditemukan larva macrobenthos lainnya, yaitu 2 jenis dari Cerithum egenum (Gould, 1849), dan Calcarina defranci d'Orbigny, 1826, 2 family/genus, yaitu: Canthocamptidae dan Portunidae, dan 2 klass, yaitu : Polychaeta dan Demospongae. Teridentifikasi jenis larva makro benthos yang baru menempel pada substrat substrat organik dan non-organik dengan ukuran yang bervariasi, yaitu dari 2 mm – 1 cm. Kepadatan jenis macrobenthos yang baru menempel adalah dari 0.03 – 0.5 individu/cm2. Ada 7 jenis teridentifkasi di lokasi Tiwoho, dan 3 jenis teridentifkasi di Malalayang Kata kunci: Penempelan, Substrat, Pesisir Tiwoho, Pesisir Malalayang

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Durairaj, Pradeepraj, Linbing Fan, Sangeeta Shrestha Sharma, Zhao Jie e Matthias Bureik. "Identification of new probe substrates for human CYP20A1". Biological Chemistry 401, n. 3 (25 febbraio 2020): 361–65. http://dx.doi.org/10.1515/hsz-2019-0307.

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AbstractCYP20A1 is a well-conserved member of the human cytochrome P450 enzyme family for which no endogenous or xenobiotic substrate is known. We have recently shown that this enzyme has moderate activity towards two proluciferin probe substrates. In order to facilitate the search for physiological substrates we have tested nine additional proluciferins in this study and identified three such probe substrates that give much higher product yields. Using one of these probes, we demonstrate inhibition of CYP20A1 activity by 1-benzylimidazole, ketoconazole and letrozole. Finally, we show that the combination of two common single nucleotide polymorphisms (SNPs) of CYP20A1 leads to an enzyme (CYP20A1Leu97Phe346) with reduced activity.

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Kleerebezem, R., e M. C. M. Van Loosdrecht. "Waste characterization for implementation in ADM1". Water Science and Technology 54, n. 4 (1 agosto 2006): 167–74. http://dx.doi.org/10.2166/wst.2006.538.

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Wastewater characterization as required for implementation in ADM1 is based on the identification of the numerous concentrations of the specific compounds defined in ADM1. However, identification of the individual substrate concentrations requires specific analytical techniques and in most cases only general measurements like COD, TOC, and organic nitrogen are available. This paper describes a simple method for calculation of the lumped elemental composition of the organic substrates in the wastewater from a limited number of widely available analyses. Using the elemental composition of the lumped substrate and the elemental composition of the substrates defined in the model, the influent composition as required for input in ADM1 can be calculated. Furthermore, proper waste characterization allows for an initial analysis of the biogas flow rate and composition as well as the reactor pH that can be achieved upon organic substrate degradation, as will be demonstrated. It is hoped that the methods described in this paper will stimulate and simplify future application of ADM1.

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Zhou, Jie, Shantao Li, Kevin K. Leung, Brian O’Donovan, James Y. Zou, Joseph L. DeRisi e James A. Wells. "Deep profiling of protease substrate specificity enabled by dual random and scanned human proteome substrate phage libraries". Proceedings of the National Academy of Sciences 117, n. 41 (24 settembre 2020): 25464–75. http://dx.doi.org/10.1073/pnas.2009279117.

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Proteolysis is a major posttranslational regulator of biology inside and outside of cells. Broad identification of optimal cleavage sites and natural substrates of proteases is critical for drug discovery and to understand protease biology. Here, we present a method that employs two genetically encoded substrate phage display libraries coupled with next generation sequencing (SPD-NGS) that allows up to 10,000-fold deeper sequence coverage of the typical six- to eight-residue protease cleavage sites compared to state-of-the-art synthetic peptide libraries or proteomics. We applied SPD-NGS to two classes of proteases, the intracellular caspases, and the ectodomains of the sheddases, ADAMs 10 and 17. The first library (Lib 10AA) allowed us to identify 104to 105unique cleavage sites over a 1,000-fold dynamic range of NGS counts and produced consensus and optimal cleavage motifs based position-specific scoring matrices. A second SPD-NGS library (Lib hP), which displayed virtually the entire human proteome tiled in contiguous 49 amino acid sequences with 25 amino acid overlaps, enabled us to identify candidate human proteome sequences. We identified up to 104natural linear cut sites, depending on the protease, and captured most of the examples previously identified by proteomics and predicted 10- to 100-fold more. Structural bioinformatics was used to facilitate the identification of candidate natural protein substrates. SPD-NGS is rapid, reproducible, simple to perform and analyze, inexpensive, and renewable, with unprecedented depth of coverage for substrate sequences, and is an important tool for protease biologists interested in protease specificity for specific assays and inhibitors and to facilitate identification of natural protein substrates.

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Kumar, Ganesan Senthil, Meng S. Choy, Dorothy M. Koveal, Michael K. Lorinsky, Scott P. Lyons, Arminja N. Kettenbach, Rebecca Page e Wolfgang Peti. "Identification of the substrate recruitment mechanism of the muscle glycogen protein phosphatase 1 holoenzyme". Science Advances 4, n. 11 (novembre 2018): eaau6044. http://dx.doi.org/10.1126/sciadv.aau6044.

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Glycogen is the primary storage form of glucose. Glycogen synthesis and breakdown are tightly controlled by glycogen synthase (GYS) and phosphorylase, respectively. The enzyme responsible for dephosphorylating GYS and phosphorylase, which results in their activation (GYS) or inactivation (phosphorylase) to robustly stimulate glycogen synthesis, is protein phosphatase 1 (PP1). However, our understanding of how PP1 recruits these substrates is limited. Here, we show how PP1, together with its muscle glycogen–targeting (GM) regulatory subunit, recruits and selectively dephosphorylates its substrates. Our molecular data reveal that the GM carbohydrate binding module (GMCBM21), which is amino-terminal to the GM PP1 binding domain, has a dual function in directing PP1 substrate specificity: It either directly recruits substrates (i.e., GYS) or recruits them indirectly by localization (via glycogen for phosphorylase). Our data provide the molecular basis for PP1 regulation by GM and reveal how PP1-mediated dephosphorylation is driven by scaffolding-based substrate recruitment.

25

Amano, Mutsuki, Yoko Kanazawa, Kei Kozawa e Kozo Kaibuchi. "Identification of the Kinase-Substrate Recognition Interface between MYPT1 and Rho-Kinase". Biomolecules 12, n. 2 (18 gennaio 2022): 159. http://dx.doi.org/10.3390/biom12020159.

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Protein kinases exert physiological functions through phosphorylating their specific substrates; however, the mode of kinase–substrate recognition is not fully understood. Rho-kinase is a Ser/Thr protein kinase that regulates cytoskeletal reorganization through phosphorylating myosin light chain (MLC) and the myosin phosphatase targeting subunit 1 (MYPT1) of MLC phosphatase (MLCP) and is involved in various diseases, due to its aberrant cellular contraction, morphology, and movement. Despite the importance of the prediction and identification of substrates and phosphorylation sites, understanding of the precise regularity in phosphorylation preference of Rho-kinase remains far from satisfactory. Here we analyzed the Rho-kinase–MYPT1 interaction, to understand the mode of Rho-kinase substrate recognition and found that the three short regions of MYPT1 close to phosphorylation sites (referred to as docking motifs (DMs); DM1 (DLQEAEKTIGRS), DM2 (KSQPKSIRERRRPR), and DM3 (RKARSRQAR)) are important for interactions with Rho-kinase. The phosphorylation levels of MYPT1 without DMs were reduced, and the effects were limited to the neighboring phosphorylation sites. We further demonstrated that the combination of pseudosubstrate (PS) and DM of MYPT1 (PS1 + DM3 and PS2 + DM2) serves as a potent inhibitor of Rho-kinase. The present information will be useful in identifying new substrates and developing selective Rho-kinase inhibitors.

26

Petrera, Agnese, Beat Amstutz, Magda Gioia, Janine Hähnlein, Antonio Baici, Petra Selchow, Davide M. Ferraris et al. "Functional characterization of the Mycobacterium tuberculosis zinc metallopeptidase Zmp1 and identification of potential substrates". Biological Chemistry 393, n. 7 (1 luglio 2012): 631–40. http://dx.doi.org/10.1515/hsz-2012-0106.

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Abstract Zinc metallopeptidases of bacterial pathogens are widely distributed virulence factors and represent promising pharmacological targets. In this work, we have characterized Zmp1, a zinc metallopeptidase identified as a virulence factor of Mycobacterium tuberculosis and belonging to the neprilysin (NEP; M13) family, whose X-ray structure has been recently solved. Interestingly, this enzyme shows an optimum activity toward a fluorogenic substrate at moderately acidic pH values (i.e., 6.3), which corresponds to those reported for the Mtb phagosome where this enzyme should exert its pathological activity. Substrate specificity of Zmp1 was investigated by screening a peptide library. Several sequences derived from biologically relevant proteins were identified as possible substrates, including the neuropeptides bradykinin, neurotensin, and neuropeptide FF. Further, subsequences of other small bioactive peptides were found among most frequently cleaved sites, e.g., apelin-13 and substance P. We determined the specific cleavage site within neuropeptides by mass spectrometry, observing that hydrophobic amino acids, mainly phenylalanine and isoleucine, are overrepresented at position P1′. In addition, the enzymatic mechanism of Zmp1 toward these neuropeptides has been characterized, displaying some differences with respect to the synthetic fluorogenic substrate and indicating that the enzyme adapts its enzymatic action to different substrates.

27

Hasyiati, Rasma, Muhammad Ali Sarong, Safrida Safrida, Djufri Djufri e Ismul Huda. "Distribution pattern of benthos based on substrate in the mangrove area of Labuhan Haji District, South Aceh Regency". Depik 12, n. 3 (26 dicembre 2023): 308–13. http://dx.doi.org/10.13170/depik.12.3.31503.

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Mangrove areas can function as habitats for spawning, rearing, and foraging for various species in them. Benthos is an organism that lives on the bottom of the water. Research on benthos distribution patterns based on substrate was carried out with the aim of studying benthos species, types of substrates, and analyzing benthos distribution patterns based on substrates in Labuhan Haji District, South Aceh District, Aceh Province. The method used in this study is the method of observation and field work. The benthos distribution pattern based on the substrate was calculated using the morphic index. Identification results found 31 species of benthos consisting of 3 classes, namely gastropods with 7 orders, bivalves with 4 orders, and malacostraca with 1 order and the type of substrate analyzed obtained 4 types of substrate namely dusty loam, sandy loam, silt and sand. The benthos distribution pattern based on the substrate obtained 2 categories, namely uniform on sandy loam and sandy clay substrates, and clustered on dusty and dusty clay substrates.Keywords:MangrovesBenthosSubstrate TypeSpread patternLabuhan Haji

28

Demir, Fatih, Stefan Niedermaier, Joji Grace Villamor e Pitter Florian Huesgen. "Quantitative proteomics in plant protease substrate identification". New Phytologist 218, n. 3 (11 maggio 2017): 936–43. http://dx.doi.org/10.1111/nph.14587.

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Kusevic, Denis, Srikanth Kudithipudi e Albert Jeltsch. "Substrate Specificity of the HEMK2 Protein Glutamine Methyltransferase and Identification of Novel Substrates". Journal of Biological Chemistry 291, n. 12 (21 gennaio 2016): 6124–33. http://dx.doi.org/10.1074/jbc.m115.711952.

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30

Nishioka, Tomoki, Mutsuki Amano, Yasuhiro Funahashi, Daisuke Tsuboi, Yukie Yamahashi e Kozo Kaibuchi. "In Vivo Identification of Protein Kinase Substrates by Kinase-Oriented Substrate Screening (KIOSS)". Current Protocols in Chemical Biology 11, n. 1 (7 gennaio 2019): e60. http://dx.doi.org/10.1002/cpch.60.

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31

Songyang, Z., K. P. Lu, Y. T. Kwon, L. H. Tsai, O. Filhol, C. Cochet, D. A. Brickey et al. "A structural basis for substrate specificities of protein Ser/Thr kinases: primary sequence preference of casein kinases I and II, NIMA, phosphorylase kinase, calmodulin-dependent kinase II, CDK5, and Erk1." Molecular and Cellular Biology 16, n. 11 (novembre 1996): 6486–93. http://dx.doi.org/10.1128/mcb.16.11.6486.

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We have developed a method to study the primary sequence specificities of protein kinases by using an oriented degenerate peptide library. We report here the substrate specificities of eight protein Ser/Thr kinases. All of the kinases studied selected distinct optimal substrates. The identified substrate specificities of these kinases, together with known crystal structures of protein kinase A, CDK2, Erk2, twitchin, and casein kinase I, provide a structural basis for the substrate recognition of protein Ser/Thr kinases. In particular, the specific selection of amino acids at the +1 and -3 positions to the substrate serine/threonine can be rationalized on the basis of sequences of protein kinases. The identification of optimal peptide substrates of CDK5, casein kinases I and II, NIMA, calmodulin-dependent kinases, Erk1, and phosphorylase kinase makes it possible to predict the potential in vivo targets of these kinases.

32

Chichkova, Nina V., Raisa A. Galiullina, Larisa V. Mochalova, Svetlana V. Trusova, Zulfazli M. Sobri, Patrick Gallois e Andrey B. Vartapetian. "Arabidopsis thaliana phytaspase: identification and peculiar properties". Functional Plant Biology 45, n. 2 (2018): 171. http://dx.doi.org/10.1071/fp16321.

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Phytaspases are plant cell death-related proteases of the subtilisin-like protease family that possess an unusual aspartate cleavage specificity. Although phytaspase activity is widespread in plants, phytaspase of Arabidopsis thaliana (L.) Heynh. has escaped detection and identification thus far. Here, we show that a single gene (At4 g10540) out of 56 A. thaliana subtilisin-like protease genes encodes a phytaspase. The recombinant phytaspase was overproduced in Nicotiana benthamiana Domin leaves, isolated, and its substrate specificity and properties were characterised. At pH 5.5, at physiological mildly acidic reaction conditions, the Arabidopsis phytaspase was shown to be strictly Asp-specific. The strongly preferred cleavage motifs of the enzyme out of a panel of synthetic peptide substrates were YVAD and IETD, while the VEID-based substrate preferred by the tobacco and rice phytaspases was almost completely resistant to hydrolysis. At neutral pH, however, the Arabidopsis phytaspase could hydrolyse peptide substrates after two additional amino acid residues, His and Phe, in addition to Asp. This observation may indicate that the repertoire of Arabidopsis phytaspase targets could possibly be regulated by the conditions of the cellular environment. Similar to tobacco and rice phytaspases, the Arabidopsis enzyme was shown to accumulate in the apoplast of epidermal leaf cells. However, in stomatal cells Arabidopsis phytaspase was observed inside the cells, possibly co-localising with vacuole. Our study thus demonstrates that the Arabidopsis phytaspase possesses both important similarities with and distinctions from the already known phytaspases, and is likely to be the most divergent member of the phytaspase family.

33

Radi, Mohammad S., Lachlan J. Munro, Daniela Rago e Douglas B. Kell. "An Untargeted Metabolomics Strategy to Identify Substrates of Known and Orphan E. coli Transporters". Membranes 14, n. 3 (20 marzo 2024): 70. http://dx.doi.org/10.3390/membranes14030070.

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Transport systems play a pivotal role in bacterial physiology and represent potential targets for medical and biotechnological applications. However, even in well-studied organisms like Escherichia coli, a notable proportion of transporters, exceeding as many as 30%, remain classified as orphans due to their lack of known substrates. This study leveraged high-resolution LC-MS-based untargeted metabolomics to identify candidate substrates for these orphan transporters. Human serum, including a diverse array of biologically relevant molecules, served as an unbiased source for substrate exposure. The analysis encompassed 26 paired transporter mutant contrasts (i.e., knockout vs. overexpression), compared with the wild type, revealing distinct patterns of substrate uptake and excretion across various mutants. The convergence of candidate substrates across mutant scenarios provided robust validation, shedding light on novel transporter-substrate relationships, including those involving yeaV, hsrA, ydjE, and yddA. Furthermore, several substrates were contingent upon the specific mutants employed. This investigation underscores the utility of untargeted metabolomics for substrate identification in the absence of prior knowledge and lays the groundwork for subsequent validation experiments, holding significant implications for both medical and biotechnological advancements.

34

Elma Eldiana, Ft Dea, Desti Christian Cahyaningrum e Bowo Nurcahyo. "Keberhasilan Identifikasi Sampel Darah Kering yang Dipaparkan pada Beragam Jenis Substrat Kayu dengan Kondisi Lingkungan Berbeda Selama Kurun Waktu Tertentu". Jurnal Biologi Indonesia 18, n. 1 (2022): 31–40. http://dx.doi.org/10.47349/jbi/18012022/31.

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Most of the criminal cases leave traces at the TKP (Tempat Kejadian Perkara) which can be used as evidence to reveal the culprit and the chronology of events that have occurred. Blood is the most important biological evidence in TKP. Through experimental research with a completely randomized design (CRD), this study aims to determine the success rate of blood groups identification in different types of Sengon (Paraserianthes falcataria L. Nielsen) as wood substrate in open and closed environmental conditions up to a certain time period. The four variations of Sengon wood substrate used were natural wood, processed wood, painted wood, and varnished wood. After 336 hours exposure on wood substrate, blood sample on the substrate was confirmed to identify the ABO system blood group using the absorption-elusion method. The results showed that the success rate identification the blood groups in ABO system on all types of Sengon wood substrate in closed environmental conditions reached a 100% for observation periods until 336 hours. Despite, the success rate of blood identification from all types wood substrate in open environmental condition were 0% for observation periods until 336 hours. Those data was then tested for normality with SPSS. The results showed that the data are not normally distributed (sig >0,00) for all type of treatmment. So that non-parametric statistical tests were carried out. The results of the Kruskall Wallis test showed that there was no significant difference between the treatment of Sengon wood substrate types and also in the length of time of observation (sig value 1,000). While the significance value on the environmental condition variable is 0.000, so it was concluded that there was a significant success rate's differences of blood group identification in samples that exposed to an open environment compared to a closed environment. This is reinforced by the results of the Kendall's correlation test and Spearman's test which show a strong correlation between the percentage of successful identification of blood groups and environmental conditions (correlation of 1,000). While the correlation value between the percentage of successful identification of blood group with the type of substrate and the length of time of exposure are 0.000 and 0.000, which is indicates a very weak correlation between the percentage of success of blood group with the type of substrate and the length of time of exposure. It can be concluded that the four types of wood substrates in closed environmental conditions had the same good ability to preserve blood within an exposure time of up to 336 hours. But, sample exposure in a closed environment provides the best percentage of success rate identification the blood groups in ABO system rather than the open environment was.

35

Elma Eldiana, Ft Dea, Desti Christian Cahyaningrum e Bowo Nurcahyo. "Keberhasilan Identifikasi Sampel Darah Kering yang Dipaparkan pada Beragam Jenis Substrat Kayu dengan Kondisi Lingkungan Berbeda Selama Kurun Waktu Tertentu". Jurnal Biologi Indonesia 18, n. 1 (2022): 31–40. http://dx.doi.org/10.47349/jbi/18012022/31.

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Abstract (sommario):

Most of the criminal cases leave traces at the TKP (Tempat Kejadian Perkara) which can be used as evidence to reveal the culprit and the chronology of events that have occurred. Blood is the most important biological evidence in TKP. Through experimental research with a completely randomized design (CRD), this study aims to determine the success rate of blood groups identification in different types of Sengon (Paraserianthes falcataria L. Nielsen) as wood substrate in open and closed environmental conditions up to a certain time period. The four variations of Sengon wood substrate used were natural wood, processed wood, painted wood, and varnished wood. After 336 hours exposure on wood substrate, blood sample on the substrate was confirmed to identify the ABO system blood group using the absorption-elusion method. The results showed that the success rate identification the blood groups in ABO system on all types of Sengon wood substrate in closed environmental conditions reached a 100% for observation periods until 336 hours. Despite, the success rate of blood identification from all types wood substrate in open environmental condition were 0% for observation periods until 336 hours. Those data was then tested for normality with SPSS. The results showed that the data are not normally distributed (sig >0,00) for all type of treatmment. So that non-parametric statistical tests were carried out. The results of the Kruskall Wallis test showed that there was no significant difference between the treatment of Sengon wood substrate types and also in the length of time of observation (sig value 1,000). While the significance value on the environmental condition variable is 0.000, so it was concluded that there was a significant success rate's differences of blood group identification in samples that exposed to an open environment compared to a closed environment. This is reinforced by the results of the Kendall's correlation test and Spearman's test which show a strong correlation between the percentage of successful identification of blood groups and environmental conditions (correlation of 1,000). While the correlation value between the percentage of successful identification of blood group with the type of substrate and the length of time of exposure are 0.000 and 0.000, which is indicates a very weak correlation between the percentage of success of blood group with the type of substrate and the length of time of exposure. It can be concluded that the four types of wood substrates in closed environmental conditions had the same good ability to preserve blood within an exposure time of up to 336 hours. But, sample exposure in a closed environment provides the best percentage of success rate identification the blood groups in ABO system rather than the open environment was.

36

Baum, Ellen Z., Steven M. Crespo-Carbone, Darren Abbanat, Barbara Foleno, Amy Maden, Raul Goldschmidt e Karen Bush. "Utility of Muropeptide Ligase for Identification of Inhibitors of the Cell Wall Biosynthesis Enzyme MurF". Antimicrobial Agents and Chemotherapy 50, n. 1 (gennaio 2006): 230–36. http://dx.doi.org/10.1128/aac.50.1.230-236.2006.

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ABSTRACT MurF is a key enzyme in the biosynthesis of the bacterial cell wall in both gram-positive and gram-negative bacteria. This enzyme has not been extensively exploited as a drug target, possibly due to the difficulty in obtaining one of the substrates, UDP-MurNAc-l-Ala-γ-d-Glu-meso-diaminopimelate, which is usually purified from bacteria. We have identified putative inhibitors of Escherichia coli MurF by a binding assay, thus bypassing the need for substrate. Inhibition of enzymatic activity was demonstrated in a high-performance liquid chromatography-based secondary assay with UDP-MurNAc-l-Ala-γ-d-Glu-diaminopimelate substrate prepared in a novel way by using muropeptide ligase enzyme to add UDP-MurNAc to synthetic l-Ala-γ-d-Glu-diaminopimelate; the substrate specificity of muropeptide ligase for peptides containing l-Lys in place of diaminopimelate was also investigated. Using the muropeptide ligase-generated MurF substrate, a thiazolylaminopyrimidine series of MurF enzyme inhibitors with 50% inhibitory concentration values as low as 2.5 μM was identified.

37

Falcocchio, Serena, Cristian Ruiz, F. I. Javier Pastor, Luciano Saso e Pilar Diaz. "Identification of a carboxylesterase-producingRhodococcussoil isolate". Canadian Journal of Microbiology 51, n. 9 (1 settembre 2005): 753–58. http://dx.doi.org/10.1139/w05-059.

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Subtropical soil microbial isolates were screened for carbohydrate, tributyrin, or olive oil hydrolysis using agar plates supplemented with the corresponding substrates. A heterotrophic, aerobic, Gram-positive strain displaying activity on tributyrin was selected and further characterized. Analysis of the morphological and physiological traits of the strain placed it as a member of the genus Rhodococcus. Further 16S rDNA sequencing revealed a 99% identity to Rhodococcus erythropolis. The strain displayed lipolytic activity on fatty-acid-derivative substrates of short chain length, with cell extract fractions having highest activity, as confirmed by the presence, after zymogram analysis, of a ca. 60-kDa intracellular protein band with activity on 4-methylumbelliferone–butyrate substrate. The presence of such a lipolytic enzyme, similar to those found in other Gram-positive bacteria, indicates that the strain could be of interest for certain biotechnological applications, like the synthesis of pharmaceuticals or biocide detoxification.Key words: Rhodococcus, lipase, esterase, soil, actinomycete.

38

Folikumah, Makafui Y., Marc Behl e Andreas Lendlein. "Reaction behaviour of peptide-based single thiol-thioesters exchange reaction substrate in the presence of externally added thiols". MRS Communications 11, n. 4 (14 luglio 2021): 402–10. http://dx.doi.org/10.1557/s43579-021-00041-z.

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Abstract Identification of patterns in chemical reaction pathways aids in the effective design of molecules for specific applications. Here, we report on model reactions with a water-soluble single thiol-thioester exchange (TTE) reaction substrate, which was designed taking in view biological and medical applications. This substrate consists of the thio-depsipeptide, Ac-Pro-Leu-Gly-SLeu-Leu-Gly-NEtSH (TDP) and does not yield foul-smelling thiol exchange products when compared with aromatic thiol containing single TTE substrates. TDP generates an α,ω-dithiol crosslinker in situ in a ‘pseudo intramolecular’ TTE. Competitive intermolecular TTE of TDP with externally added “basic” thiols increased the crosslinker concentration whilst “acidic” thiols decreased its concentration. TDP could potentially enable in situ bioconjugation and crosslinking applications. Graphic abstract The competition between ‘pseudo intramolecular’ and intermolecular exchange of a peptide-based thiol-thioester exchange (TTE) substrate can be used to control the relative amount of final exchange products based on size and pKa values of externally added thiols. Potential application of this system can be seen in the development of TTE substrates for the rapid identification of thiols by dynamic combinatorial screening.

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McAdam, Steven O. "Effects of substrate condition on habitat use and survival by white sturgeon (Acipenser transmontanus) larvae and potential implications for recruitment". Canadian Journal of Fisheries and Aquatic Sciences 68, n. 5 (maggio 2011): 812–22. http://dx.doi.org/10.1139/f2011-021.

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To understand links between substrate and recruitment of white sturgeon ( Acipenser transmontanus ), I evaluated the effects of substrate condition on larval drift, hiding, and predation between hatch and 15 days posthatch (dph). Over porous substrates (small gravel = 1.2–1.9 cm; medium gravel = 2.5–5.0 cm; cobble = 10–15 cm), rapid interstitial hiding was observed from 0 to 6 dph at low water velocity (4 cm·s–1), whereas larvae drifted in response to nonporous substrates (sand < 0.2 cm; embedded cobble). Velocities of 20 cm·s–1 led to significantly lower drift only at 1 dph over small gravel. Hiding occurred an average of 2.0–13.3 s after release at 0–6 dph. Predation by sculpins ( Cottus spp.) on larval sturgeon also decreased significantly in response to porous substrates at 1 dph. The strongest expression of increased hiding and decreased predation when small gravel was available suggests that yolksac larvae prefer small interstitial spaces created by that substrate. Considering behavioural responses in preferred natural spawning habitat suggests yolksac larvae predominantly hide in the vicinity of spawning locations. Identification of strong effects of substrate condition on age-specific drift and survival suggests that substrate degradation may contribute to recruitment limitations for sturgeon.

40

Menolli Junior, Nelson, Tatiane Asai, Marina Capelari e Luzia Doretto Paccola-Meirelles. "Morphological and molecular identification of four Brazilian commercial isolates of Pleurotus spp. and cultivation on corncob". Brazilian Archives of Biology and Technology 53, n. 2 (aprile 2010): 397–408. http://dx.doi.org/10.1590/s1516-89132010000200019.

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The species of Pleurotus have great commercial importance and adaptability for growth and fructification within a wide variety of agro-industrial lignocellulosic wastes. In this study, two substrates prepared from ground corncobs supplemented with rice bran and charcoal were tested for mycelium growth kinetics in test tubes and for the cultivation of four Pleurotus commercial isolates in polypropylene bags. The identification of the isolates was based on the morphology of the basidiomata obtained and on sequencing of the LSU rDNA gene. Three isolates were identified as P. ostreatus, and one was identified as P. djamor. All isolates had better in-depth mycelium development in the charcoal-supplemented substrate. In the cultivation experiment, the isolates reacted differently to the two substrates. One isolate showed particularly high growth on the substrate containing charcoal.

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Dott, W., e P. Kämpfer. "Biochemical Methods for Automated Bacterial Identification and Testing Metabolic Activities in Water and Wastewater". Water Science and Technology 20, n. 11-12 (1 novembre 1988): 221–27. http://dx.doi.org/10.2166/wst.1988.0288.

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The improvements of computer-assisted identification were used to develop a new microplate system for characterization and identification of various gram-negative and gram-positive heterotrophic bacteria from the environment. In standard microtitration plates about 90 biochemical tests, some of them conventional tests, carbon substrate assimilation tests and enzyme tests using chromogenic substrates can be performed. Reading of the test results is done automatically by a photometer coupled to a computer. The applications of this identification system are shown by two waste water samples.

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Newton, Philip M., e Robert O. Messing. "The substrates and binding partners of protein kinase Cε". Biochemical Journal 427, n. 2 (29 marzo 2010): 189–96. http://dx.doi.org/10.1042/bj20091302.

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The ε isoform of protein kinase C (PKCε) has important roles in the function of the cardiac, immune and nervous systems. As a result of its diverse actions, PKCε is the target of active drug-discovery programmes. A major research focus is to identify signalling cascades that include PKCε and the substrates that PKCε regulates. In the present review, we identify and discuss those proteins that have been conclusively shown to be direct substrates of PKCε by the best currently available means. We will also describe binding partners that anchor PKCε near its substrates. We review the consequences of substrate phosphorylation and discuss cellular mechanisms by which target specificity is achieved. We begin with a brief overview of the biology of PKCε and methods for substrate identification, and proceed with a discussion of substrate categories to identify common themes that emerge and how these may be used to guide future studies.

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Koudelakova, Tana, Eva Chovancova, Jan Brezovsky, Marta Monincova, Andrea Fortova, Jiri Jarkovsky e Jiri Damborsky. "Substrate specificity of haloalkane dehalogenases". Biochemical Journal 435, n. 2 (29 marzo 2011): 345–54. http://dx.doi.org/10.1042/bj20101405.

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An enzyme's substrate specificity is one of its most important characteristics. The quantitative comparison of broad-specificity enzymes requires the selection of a homogenous set of substrates for experimental testing, determination of substrate-specificity data and analysis using multivariate statistics. We describe a systematic analysis of the substrate specificities of nine wild-type and four engineered haloalkane dehalogenases. The enzymes were characterized experimentally using a set of 30 substrates selected using statistical experimental design from a set of nearly 200 halogenated compounds. Analysis of the activity data showed that the most universally useful substrates in the assessment of haloalkane dehalogenase activity are 1-bromobutane, 1-iodopropane, 1-iodobutane, 1,2-dibromoethane and 4-bromobutanenitrile. Functional relationships among the enzymes were explored using principal component analysis. Analysis of the untransformed specific activity data revealed that the overall activity of wild-type haloalkane dehalogenases decreases in the following order: LinB~DbjA>DhlA~DhaA~DbeA~DmbA>DatA~DmbC~DrbA. After transforming the data, we were able to classify haloalkane dehalogenases into four SSGs (substrate-specificity groups). These functional groups are clearly distinct from the evolutionary subfamilies, suggesting that phylogenetic analysis cannot be used to predict the substrate specificity of individual haloalkane dehalogenases. Structural and functional comparisons of wild-type and mutant enzymes revealed that the architecture of the active site and the main access tunnel significantly influences the substrate specificity of these enzymes, but is not its only determinant. The identification of other structural determinants of the substrate specificity remains a challenge for further research on haloalkane dehalogenases.

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Chapelat, Julien, Frédéric Berst, Andreas L. Marzinzik, Henrik Moebitz, Peter Drueckes, Doriano Fabbro, Joerg Trappe e Dieter Seebach. "Substrate profiling of IGF-1R and InsR: Identification of a potent pentamer substrate". Bioorganic & Medicinal Chemistry Letters 21, n. 23 (dicembre 2011): 7030–33. http://dx.doi.org/10.1016/j.bmcl.2011.09.101.

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Fang, Mingyu, Xing Wang, Zhikun Jia, Qiongju Qiu, Peng Li, Li Chen e Hui Yang. "A Simple and Efficient Method for the Substrate Identification of Amino Acid Decarboxylases". International Journal of Molecular Sciences 23, n. 23 (22 novembre 2022): 14551. http://dx.doi.org/10.3390/ijms232314551.

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Abstract (sommario):

Amino acid decarboxylases convert amino acids into different biogenic amines which regulate diverse biological processes. Therefore, identifying the substrates of amino acid decarboxylases is critical for investigating the function of the decarboxylases, especially for the new genes predicted to be amino acid decarboxylases. In the present work, we have established a simple and efficient method to identify the substrates and enzymatic activity of amino acid decarboxylases based on LC-MS methods. We chose GAD65 and AADC as models to validate our method. GAD65 and AADC were expressed in HEK 293T cells and purified through immunoprecipitation. The purified amino acid decarboxylases were subjected to enzymatic reaction with different substrate mixtures in vitro. LC-MS analysis of the reaction mixture identified depleted or accumulated metabolites, which corresponded to candidate enzyme substrates and products, respectively. Our method successfully identified the substrates and products of known amino acid decarboxylases. In summary, our method can efficiently identify the substrates and products of amino acid decarboxylases, which will facilitate future amino acid decarboxylase studies.

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Ibrahim, Mohamad Mokhtar, Zulkifly Jemaat e Abdurahman Hamid Nour. "Microbiota of a UASB Reactor Treating Palm Oil Mill Effluent Using HiSeq Sequencing". Materials Science Forum 1025 (marzo 2021): 169–76. http://dx.doi.org/10.4028/www.scientific.net/msf.1025.169.

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Potential microbial communities in the UASB reactor fed with two different substrates i.e synthetic wastewater and raw palm oil mill effluent (POME) were elucidated by using one of the most popular techniques in molecular methods, viz 16S rDNA cloning. The methodology involved; the extraction of nucleic acids, amplification and cloning of the 16S rRNA genes on sequencing HiSeq platform and finally identification and affiliation of the isolated clone with the aid of phylogenetic software. Results showed that the genus methanosarcina and methanosaeta were dominant methanogens in this study for both substrates types. Overall, microbial population (Bacteria and Archaea) in sample A (POME as substrate) is more diverse compared to sample B (synthetic wastewater as substrate) due to abundance of microorganism population in raw POME which was used as a substrate. However for the methanogenic (Archaea) diversity in both samples, there was not much different between sample A and sample B probably due to similar inoculum was inoculated in the reactor despite of have using different substrate type.

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Smirnova, Maria, Laura Goracci, Gabriele Cruciani, Laetitia Federici, Xavier Declèves, Hélène Chapy e Salvatore Cisternino. "Pharmacophore-Based Discovery of Substrates of a Novel Drug/Proton-Antiporter in the Human Brain Endothelial hCMEC/D3 Cell Line". Pharmaceutics 14, n. 2 (21 gennaio 2022): 255. http://dx.doi.org/10.3390/pharmaceutics14020255.

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A drug/proton-antiporter, whose the molecular structure is still unknown, was previously evidenced at the blood-brain barrier (BBB) by functional experiments. The computational method could help in the identification of substrates of this solute carrier (SLC) transporter. Two pharmacophore models for substrates of this transporter using the FLAPpharm approach were developed. The trans-stimulation potency of 40 selected compounds for already known specific substrates ([3H]-clonidine) were determined and compared in the human brain endothelial cell line hCMEC/D3. Results. The two pharmacophore models obtained were used as templates to screen xenobiotic and endogenous compounds from four databases (e.g., Specs), and 45 hypothetical new candidates were tested to determine their substrate capacity. Psychoactive drugs such as antidepressants (e.g., imipramine, desipramine), antipsychotics/neuroleptics such as phenothiazine derivatives (chlorpromazine), sedatives anti-histamine-H1 drugs (promazine, promethazine, triprolidine, pheniramine), opiates/opioids (e.g., hydrocodone), trihexyphenidyl and sibutramine were correctly predicted as proton-antiporter substrates. The best performing pharmacophore model for the proton-antiporter substrates appeared as a good predictor of known substrates and allowed the identification of new substrate compounds. This model marks a new step in the characterization of this drug/proton-antiporter and will be of great use in uncovering its substrates and designing chemical entities with an improved influx capability to cross the BBB.

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DeMarco, Andrew G., e Mark C. Hall. "Phosphoproteomic Approaches for Identifying Phosphatase and Kinase Substrates". Molecules 28, n. 9 (24 aprile 2023): 3675. http://dx.doi.org/10.3390/molecules28093675.

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Protein phosphorylation is a ubiquitous post-translational modification controlled by the opposing activities of protein kinases and phosphatases, which regulate diverse biological processes in all kingdoms of life. One of the key challenges to a complete understanding of phosphoregulatory networks is the unambiguous identification of kinase and phosphatase substrates. Liquid chromatography-coupled mass spectrometry (LC-MS/MS) and associated phosphoproteomic tools enable global surveys of phosphoproteome changes in response to signaling events or perturbation of phosphoregulatory network components. Despite the power of LC-MS/MS, it is still challenging to directly link kinases and phosphatases to specific substrate phosphorylation sites in many experiments. Here, we survey common LC-MS/MS-based phosphoproteomic workflows for identifying protein kinase and phosphatase substrates, noting key advantages and limitations of each. We conclude by discussing the value of inducible degradation technologies coupled with phosphoproteomics as a new approach that overcomes some limitations of current methods for substrate identification of kinases, phosphatases, and other regulatory enzymes.

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Yuan, Y., e S. Altman. "Substrate recognition by human RNase P: identification of small, model substrates for the enzyme." EMBO Journal 14, n. 1 (gennaio 1995): 159–68. http://dx.doi.org/10.1002/j.1460-2075.1995.tb06986.x.

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Law, Simon, Xin Du, Preety Panwar, Nicolette S. Honson, Tom Pfeifer, Michel Roberge e Dieter Brömme. "Identification of substrate-specific inhibitors of cathepsin K through high-throughput screening". Biochemical Journal 476, n. 3 (5 febbraio 2019): 499–512. http://dx.doi.org/10.1042/bcj20180851.

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Abstract Cathepsin K (CatK) is a cysteine protease and drug target for skeletal disorders that is known for its potent collagenase and elastase activity. The formation of oligomeric complexes of CatK in the presence of glycosaminoglycans has been associated with its collagenase activity. Inhibitors that disrupt these complexes can selectively block the collagenase activity without interfering with the other regulatory proteolytic activities of the enzyme. Here, we have developed a fluorescence polarization (FP) assay to screen 4761 compounds for substrate-specific ectosteric collagenase inhibitors of CatK. A total of 38 compounds were identified that block the collagenase activity without interfering with the hydrolysis of active site substrates such as the synthetic peptide substrate, benzyloxycarbonyl-Phe-Arg-7-amido-4-methylcoumarin, and gelatin. The identified inhibitors can be divided into two main classes, negatively charged and polyaromatic compounds which suggest the binding to different ectosteric sites. Two of the inhibitors were highly effective in preventing the bone-resorption activity of CatK in osteoclasts. Interestingly, some of the ectosteric inhibitors were capable of differentiating between the collagenase and elastase activity of CatK depending on the ectosteric site utilized by the compound. Owing to their substrate-specific selectivity, ectosteric inhibitors represent a viable alternative to side effect-prone active site-directed inhibitors.

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