Journal articles: 'ADN Ligase'

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Lee, Jaeseok, Youngjun Lee, Young Mee Jung, Ju Hyun Park, Hyuk Sang Yoo, and Jongmin Park. "Discovery of E3 Ligase Ligands for Target Protein Degradation." Molecules 27, no. 19 (October 2, 2022): 6515. http://dx.doi.org/10.3390/molecules27196515.

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Target protein degradation has emerged as a promising strategy for the discovery of novel therapeutics during the last decade. Proteolysis-targeting chimera (PROTAC) harnesses a cellular ubiquitin-dependent proteolysis system for the efficient degradation of a protein of interest. PROTAC consists of a target protein ligand and an E3 ligase ligand so that it enables the target protein degradation owing to the induced proximity with ubiquitin ligases. Although a great number of PROTACs has been developed so far using previously reported ligands of proteins for their degradation, E3 ligase ligands have been mostly limited to either CRBN or VHL ligands. Those PROTACs showed their limitation due to the cell type specific expression of E3 ligases and recently reported resistance toward PROTACs with CRBN ligands or VHL ligands. To overcome these hurdles, the discovery of various E3 ligase ligands has been spotlighted to improve the current PROTAC technology. This review focuses on currently reported E3 ligase ligands and their application in the development of PROTACs.

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Tomkinson, Alan E., Tasmin Naila, and Seema Khattri Bhandari. "Altered DNA ligase activity in human disease." Mutagenesis 35, no. 1 (October 20, 2019): 51–60. http://dx.doi.org/10.1093/mutage/gez026.

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Abstract The joining of interruptions in the phosphodiester backbone of DNA is critical to maintain genome stability. These breaks, which are generated as part of normal DNA transactions, such as DNA replication, V(D)J recombination and meiotic recombination as well as directly by DNA damage or due to DNA damage removal, are ultimately sealed by one of three human DNA ligases. DNA ligases I, III and IV each function in the nucleus whereas DNA ligase III is the sole enzyme in mitochondria. While the identification of specific protein partners and the phenotypes caused either by genetic or chemical inactivation have provided insights into the cellular functions of the DNA ligases and evidence for significant functional overlap in nuclear DNA replication and repair, different results have been obtained with mouse and human cells, indicating species-specific differences in the relative contributions of the DNA ligases. Inherited mutations in the human LIG1 and LIG4 genes that result in the generation of polypeptides with partial activity have been identified as the causative factors in rare DNA ligase deficiency syndromes that share a common clinical symptom, immunodeficiency. In the case of DNA ligase IV, the immunodeficiency is due to a defect in V(D)J recombination whereas the cause of the immunodeficiency due to DNA ligase I deficiency is not known. Overexpression of each of the DNA ligases has been observed in cancers. For DNA ligase I, this reflects increased proliferation. Elevated levels of DNA ligase III indicate an increased dependence on an alternative non-homologous end-joining pathway for the repair of DNA double-strand breaks whereas elevated level of DNA ligase IV confer radioresistance due to increased repair of DNA double-strand breaks by the major non-homologous end-joining pathway. Efforts to determine the potential of DNA ligase inhibitors as cancer therapeutics are on-going in preclinical cancer models.

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Cao, Weiguo. "DNA ligases and ligase-based technologies." Clinical and Applied Immunology Reviews 2, no. 1 (November 2001): 33–43. http://dx.doi.org/10.1016/s1529-1049(01)00039-3.

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Fang, Deyu, An Chen, and Sang-Myeong Lee. "Inhibition of activation-induced T cell death by AIP2-mediated ubiquitination of EGR2 (35.20)." Journal of Immunology 182, no. 1_Supplement (April 1, 2009): 35.20. http://dx.doi.org/10.4049/jimmunol.182.supp.35.20.

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Abstract E3 ubiquitin ligases, which target specific molecules for proteolytic destruction, have emerged as key regulators of immune functions. Several E3 ubiquitin ligases, including c-Cbl, Cbl-b, GRAIL, Itch, and Nedd4, have been shown to negatively regulate T-cell activation. Here we report that the HECT-type E3 ligase, AIP2, positively regulates T-cell activation. Ectopic expression of AIP2 in mouse primary T cells enhances their proliferation and IL-2 production by suppressing apoptosis of T cells. AIP2 interacts with and promotes ubiquitin-mediated degradation of EGR2, a zinc finger transcription factor that has been found to regulate Fas ligand (FasL) expression during activation-induced T cell death. Suppression of AIP2 expression by small RNA interference upregulates EGR2 and FasL expression and enhances the apoptosis of T cells. Therefore, AIP2 regulates activation-induced T cell death by suppressing EGR2-mediated FasL expression via the ubiquitin pathway.

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Kennan, Alan J., V. Haridas, Kay Severin, David H. Lee, and M. Reza Ghadiri. "Ade NovoDesigned Peptide Ligase: A Mechanistic Investigation." Journal of the American Chemical Society 123, no. 9 (March 2001): 1797–803. http://dx.doi.org/10.1021/ja991266c.

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Fanucci, Francesco. "Quaternary shorelines and continental shelf of the Ligurian coast." Zeitschrift für Geomorphologie 31, no. 4 (December 17, 1987): 463–72. http://dx.doi.org/10.1127/zfg/31/1987/463.

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Gu, Jiafeng, Haihui Lu, Brigette Tippin, Noriko Shimazaki, Myron F. Goodman, and Michael R. Lieber. "XRCC4:DNA ligase IV can ligate incompatible DNA ends and can ligate across gaps." EMBO Journal 26, no. 14 (July 25, 2007): 3506–7. http://dx.doi.org/10.1038/sj.emboj.7601729.

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Thayale Purayil, Fayas, Naganeeswaran Sudalaimuthuasari, Ling Li, Ruwan Aljneibi, Aysha Mohammed Khamis Al Shamsi, Nelson David, Martin Kottackal, et al. "Transcriptome Profiling and Functional Validation of RING-Type E3 Ligases in Halophyte Sesuvium verrucosum under Salinity Stress." International Journal of Molecular Sciences 23, no. 5 (March 4, 2022): 2821. http://dx.doi.org/10.3390/ijms23052821.

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Owing to their sessile nature, plants have developed a tapestry of molecular and physiological mechanisms to overcome diverse environmental challenges, including abiotic stresses. Adaptive radiation in certain lineages, such as Aizoaceae, enable their success in colonizing arid regions and is driven by evolutionary selection. Sesuvium verrucosum (commonly known as Western sea-purslane) is a highly salt-tolerant succulent halophyte belonging to the Aizoaceae family; thus, it provides us with the model-platform for studying plant adaptation to salt stress. Various transcriptional and translational mechanisms are employed by plants to cope with salt stress. One of the systems, namely, ubiquitin-mediated post-translational modification, plays a vital role in plant tolerance to abiotic stress and other biological process. E3 ligase plays a central role in target recognition and protein specificity in ubiquitin-mediated protein degradation. Here, we characterize E3 ligases in Sesuvium verrucosum from transcriptome analysis of roots in response to salinity stress. Our de novo transcriptome assembly results in 131,454 transcripts, and the completeness of transcriptome was confirmed by BUSCO analysis (99.3% of predicted plant-specific ortholog genes). Positive selection analysis shows 101 gene families under selection; these families are enriched for abiotic stress (e.g., osmotic and salt) responses and proteasomal ubiquitin-dependent protein catabolic processes. In total, 433 E3 ligase transcripts were identified in S. verrucosum; among these transcripts, single RING-type classes were more abundant compared to multi-subunit RING-type E3 ligases. Additionally, we compared the number of single RING-finger E3 ligases with ten different plant species, which confirmed the abundance of single RING-type E3 ligases in different plant species. In addition, differential expression analysis showed significant changes in 13 single RING-type E3 ligases (p-value < 0.05) under salinity stress. Furthermore, the functions of the selected E3 ligases genes (12 genes) were confirmed by yeast assay. Among them, nine genes conferred salt tolerance in transgenic yeast. This functional assay supports the possible involvement of these E3 ligase in salinity stress. Our results lay a foundation for translational research in glycophytes to develop stress tolerant crops.

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Gong, Yao, and Yue Chen. "UbE3-APA: a bioinformatic strategy to elucidate ubiquitin E3 ligase activities in quantitative proteomics study." Bioinformatics 38, no. 8 (February 9, 2022): 2211–18. http://dx.doi.org/10.1093/bioinformatics/btac069.

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Abstract Motivation Ubiquitination is widely involved in protein homeostasis and cell signaling. Ubiquitin E3 ligases are critical regulators of ubiquitination that recognize and recruit specific ubiquitination targets for the final rate-limiting step of ubiquitin transfer reactions. Understanding the ubiquitin E3 ligase activities will provide knowledge in the upstream regulator of the ubiquitination pathway and reveal potential mechanisms in biological processes and disease progression. Recent advances in mass spectrometry-based proteomics have enabled deep profiling of ubiquitylome in a quantitative manner. Yet, functional analysis of ubiquitylome dynamics and pathway activity remains challenging. Results Here, we developed a UbE3-APA, a computational algorithm and stand-alone python-based software for Ub E3 ligase Activity Profiling Analysis. Combining an integrated annotation database with statistical analysis, UbE3-APA identifies significantly activated or suppressed E3 ligases based on quantitative ubiquitylome proteomics datasets. Benchmarking the software with published quantitative ubiquitylome analysis confirms the genetic manipulation of SPOP enzyme activity through overexpression and mutation. Application of the algorithm in the re-analysis of a large cohort of ubiquitination proteomics study revealed the activation of PARKIN and the co-activation of other E3 ligases in mitochondria depolarization-induced mitophagy process. We further demonstrated the application of the algorithm in the DIA (data-independent acquisition)-based quantitative ubiquitylome analysis. Availability and implementation Source code and binaries are freely available for download at URL: https://github.com/Chenlab-UMN/Ub-E3-ligase-Activity-Profiling-Analysis, implemented in python and supported on Linux and MS Windows. Supplementary information Supplementary data are available at Bioinformatics online.

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Alomari, Arqam, Robert Gowland, Callum Southwood, Jak Barrow, Zoe Bentley, Jashel Calvin-Nelson, Alice Kaminski, et al. "Identification of Novel Inhibitors of Escherichia coli DNA Ligase (LigA)." Molecules 26, no. 9 (April 25, 2021): 2508. http://dx.doi.org/10.3390/molecules26092508.

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Present in all organisms, DNA ligases catalyse the formation of a phosphodiester bond between a 3′ hydroxyl and a 5′ phosphate, a reaction that is essential for maintaining genome integrity during replication and repair. Eubacterial DNA ligases use NAD+ as a cofactor and possess low sequence and structural homology relative to eukaryotic DNA ligases which use ATP as a cofactor. These key differences enable specific targeting of bacterial DNA ligases as an antibacterial strategy. In this study, four small molecule accessible sites within functionally important regions of Escherichia coli ligase (EC-LigA) were identified using in silico methods. Molecular docking was then used to screen for small molecules predicted to bind to these sites. Eight candidate inhibitors were then screened for inhibitory activity in an in vitro ligase assay. Five of these (geneticin, chlorhexidine, glutathione (reduced), imidazolidinyl urea and 2-(aminomethyl)imidazole) showed dose-dependent inhibition of EC-LigA with half maximal inhibitory concentrations (IC50) in the micromolar to millimolar range (11–2600 µM). Two (geneticin and chlorhexidine) were predicted to bind to a region of EC-LigA that has not been directly investigated previously, raising the possibility that there may be amino acids within this region that are important for EC-LigA activity or that the function of essential residues proximal to this region are impacted by inhibitor interactions with this region. We anticipate that the identified small molecule binding sites and inhibitors could be pursued as part of an antibacterial strategy targeting bacterial DNA ligases.

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Sarri, Niki, Natalia Papadopoulos, Johan Lennartsson, and Carl-Henrik Heldin. "The E3 Ubiquitin Ligase TRIM21 Regulates Basal Levels of PDGFRβ." International Journal of Molecular Sciences 24, no. 9 (April 24, 2023): 7782. http://dx.doi.org/10.3390/ijms24097782.

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Activation of platelet-derived growth factor (PDGF) receptors α and β (PDGFRα and PDGFRβ) at the cell surface by binding of PDGF isoforms leads to internalization of receptors, which affects the amplitude and kinetics of signaling. Ubiquitination of PDGF receptors in response to ligand stimulation is mediated by the Casitas b-lineage lymphoma (Cbl) family of ubiquitin ligases, promoting internalization and serving as a sorting signal for vesicular trafficking of receptors. We report here that another E3 ligase, i.e., tripartite motif-containing protein 21 (TRIM21), contributes to the ubiquitination of PDGFRβ in human primary fibroblasts AG1523 and the osteosarcoma cell line U2OS and regulates basal levels of PDGFRβ. We found that siRNA-mediated depletion of TRIM21 led to decreased ubiquitination of PDGFRβ in response to PDGF-BB stimulation, while internalization from the cell surface and the rate of ligand-induced degradation of the receptor were not affected. Moreover, induction of TRIM21 decreased the levels of PDGFRβ in serum-starved cells, and even more in growing cells, in the absence of PDGF stimulation. Consistently, siRNA knockdown of TRIM21 caused accumulation of the total amount of PDGFRβ, both in the cytoplasm and on the cell surface, without affecting mRNA levels of the receptor. We conclude that TRIM21 acts post-translationally and maintains basal levels of PDGFRβ, thus suggesting that ubiquitination of PDGFRβ by TRIM21 may direct a portion of receptor for degradation in growing cells in a ligand-independent manner.

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Goto, Jun, Yoichiro Otaki, Tetsu Watanabe, and Masafumi Watanabe. "The Role of HECT-Type E3 Ligase in the Development of Cardiac Disease." International Journal of Molecular Sciences 22, no. 11 (June 4, 2021): 6065. http://dx.doi.org/10.3390/ijms22116065.

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Despite advances in medicine, cardiac disease remains an increasing health problem associated with a high mortality rate. Maladaptive cardiac remodeling, such as cardiac hypertrophy and fibrosis, is a risk factor for heart failure; therefore, it is critical to identify new therapeutic targets. Failing heart is reported to be associated with hyper-ubiquitylation and impairment of the ubiquitin–proteasome system, indicating an importance of ubiquitylation in the development of cardiac disease. Ubiquitylation is a post-translational modification that plays a pivotal role in protein function and degradation. In 1995, homologous to E6AP C-terminus (HECT) type E3 ligases were discovered. E3 ligases are key enzymes in ubiquitylation and are classified into three families: really interesting new genes (RING), HECT, and RING-between-RINGs (RBRs). Moreover, 28 HECT-type E3 ligases have been identified in human beings. It is well conserved in evolution and is characterized by the direct attachment of ubiquitin to substrates. HECT-type E3 ligase is reported to be involved in a wide range of human diseases and health. The role of HECT-type E3 ligases in the development of cardiac diseases has been uncovered in the last decade. There are only a few review articles summarizing recent advancements regarding HECT-type E3 ligase in the field of cardiac disease. This study focused on cardiac remodeling and described the role of HECT-type E3 ligases in the development of cardiac disease. Moreover, this study revealed that the current knowledge could be exploited for the development of new clinical therapies.

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Sharma, Chiranjeev, Myeong A. Choi, Yoojin Song, and Young Ho Seo. "Rational Design and Synthesis of HSF1-PROTACs for Anticancer Drug Development." Molecules 27, no. 5 (March 2, 2022): 1655. http://dx.doi.org/10.3390/molecules27051655.

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PROTACs employ the proteosome-mediated proteolysis via E3 ligase and recruit the natural protein degradation machinery to selectively degrade the cancerous proteins. Herein, we have designed and synthesized heterobifunctional small molecules that consist of different linkers tethering KRIBB11, a HSF1 inhibitor, with pomalidomide, a commonly used E3 ligase ligand for anticancer drug development.

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Kounde, Cyrille S., Maria M. Shchepinova, Charlie N. Saunders, Marcel Muelbaier, Mark D. Rackham, John D. Harling, and Edward W. Tate. "A caged E3 ligase ligand for PROTAC-mediated protein degradation with light." Chemical Communications 56, no. 41 (2020): 5532–35. http://dx.doi.org/10.1039/d0cc00523a.

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Wei, Wei, Jian-ye Chen, Ze-xiang Zeng, Jian-fei Kuang, Wang-jin Lu, and Wei Shan. "The Ubiquitin E3 Ligase MaLUL2 Is Involved in High Temperature-Induced Green Ripening in Banana Fruit." International Journal of Molecular Sciences 21, no. 24 (December 9, 2020): 9386. http://dx.doi.org/10.3390/ijms21249386.

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Harvested banana fruit ripened under warm temperatures above 24 °C remain green peel, leading to severe economic loss. E3 ubiquitin-ligases, as the major components in the ubiquitination pathway, have been implicated to play important roles in temperature-stress responses. However, the molecular mechanism underlying high temperature-triggered stay-green ripening bananas in association with E3 ubiquitin-ligases, remains largely unknown. In this study, a RING-type E3 ubiquitin ligase termed MaLUL2, was isolated and characterized from banana fruit. The MaLUL2 gene contains 1095 nucleotides and encodes a protein with 365 amino acids. The MaLUL2 protein contains a domain associated with RING2 (DAR2) and a RING domain, which are the typical characteristics of RING-type E3 ligases. MaLUL2 expression was up-regulated during high temperature-induced green ripening. Subcellular localization showed that MaLUL2 localized in the nucleus, cytoplasm, and plasma membrane. MaLUL2 displayed E3 ubiquitin ligase activity in vitro. More importantly, transient overexpression of MaLUL2 in banana fruit peel increased the level of ubiquitination in vivo and led to a stay-green phenotype, accompanying with decreased expression of chlorophyll catabolic genes. Collectively, these findings suggest that MaLUL2 might act as a negative regulator of chlorophyll degradation and provide novel insights into the regulatory mechanism of high temperature-induced green ripening bananas.

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Toma-Fukai, Sachiko, and Toshiyuki Shimizu. "Structural Diversity of Ubiquitin E3 Ligase." Molecules 26, no. 21 (November 4, 2021): 6682. http://dx.doi.org/10.3390/molecules26216682.

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The post-translational modification of proteins regulates many biological processes. Their dysfunction relates to diseases. Ubiquitination is one of the post-translational modifications that target lysine residue and regulate many cellular processes. Three enzymes are required for achieving the ubiquitination reaction: ubiquitin-activating enzyme (E1), ubiquitin-conjugating enzyme (E2), and ubiquitin ligase (E3). E3s play a pivotal role in selecting substrates. Many structural studies have been conducted to reveal the molecular mechanism of the ubiquitination reaction. Recently, the structure of PCAF_N, a newly categorized E3 ligase, was reported. We present a review of the recent progress toward the structural understanding of E3 ligases.

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FOGAR, Sandra R. "LA MULTIDIMENSIONALIDAD DEL ESTUDIO DEL TERRITORIO Y EL DERECHO A LA TERRITORIALIDAD DESDE UNA PERSPECTIVA DE CONSTRUCCIÓN DE TERRITORIALIDAD." ADNea, no. 8 (November 19, 2020): 132. http://dx.doi.org/10.30972/adn.084580.

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Se parte de describir la naturaleza epistemológica del concepto de territorio. Se asume que su condición de objeto de estudio complejo conduce a buscar respuestas en un campo de análisis más amplio en el que confluyen diversas disciplinas. Se aborda el territorio, escenario de múltiples problemáticas. La mirada multidimensional desde la perspectiva crítica del conocimiento y de lo social posibilita pensar la ocupación e intervención territorial, ligada a escenarios de la realidad que anuncian relaciones de dominación y resistencia, y derivado de ello, la Construcción de Territorialidad como conocimiento emancipatorio desde el cual reflexionar acerca del Derecho a la Territorialidad.

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Andrews, Paul S., Steve Schneider, Evelyn Yang, Mark Michaels, Hao Chen, Jin Tang, and Renee Emkey. "Identification of Substrates of SMURF1 Ubiquitin Ligase Activity Utilizing Protein Microarrays." ASSAY and Drug Development Technologies 8, no. 4 (August 2010): 471–87. http://dx.doi.org/10.1089/adt.2009.0264.

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Corbridge, Emily, Alexandra MacGregor, Raed Al-Saharin, Matthew G. Garneau, Samuel Smalley, Sutton Mooney, Sanja Roje, Philip D. Bates, and Hanjo Hellmann. "Brassica napus Plants Gain Improved Salt-Stress Tolerance and Increased Storage Oil Biosynthesis by Interfering with CRL3BPM Activities." Plants 12, no. 5 (March 1, 2023): 1085. http://dx.doi.org/10.3390/plants12051085.

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Generating new strategies to improve plant performance and yield in crop plants becomes increasingly relevant with ongoing and predicted global climate changes. E3 ligases that function as key regulators within the ubiquitin proteasome pathway often are involved in abiotic stress responses, development, and metabolism in plants. The aim of this research was to transiently downregulate an E3 ligase that uses BTB/POZ-MATH proteins as substrate adaptors in a tissue-specific manner. Interfering with the E3 ligase at the seedling stage and in developing seeds results in increased salt-stress tolerance and elevated fatty acid levels, respectively. This novel approach can help to improve specific traits in crop plants to maintain sustainable agriculture.

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Sun, Aiqin, Yifei Chen, Xianyan Tian, and Qiong Lin. "The Role of HECT E3 Ubiquitin Ligases in Colorectal Cancer." Biomedicines 11, no. 2 (February 7, 2023): 478. http://dx.doi.org/10.3390/biomedicines11020478.

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Colorectal cancer (CRC) is estimated to rank as the second reason for cancer-related deaths, and the prognosis of CRC patients remains unsatisfactory. Numerous studies on gastrointestinal cell biology have shown that the E3 ligase-mediated ubiquitination exerts key functions in the pathogenesis of CRC. The homologous to E6-associated protein C-terminus (HECT) family E3 ligases are a major group of E3 enzymes, featured with the presence of a catalytic HECT domain, which participate in multiple cellular processes; thus, alterations in HECT E3 ligases in function or expression are closely related to the occurrence and development of many human malignancies, including—but not limited to—CRC. In this review, we summarize the potential role of HECT E3 ligases in colorectal carcinogenesis and the related underlying molecular mechanism to expand our understanding of their pathological functions. Exploiting specific inhibitors targeting HECT E3 ligases could be a potential therapeutic strategy for CRC therapy in the future.

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Wang, Peiwen, Lin Zhu, Ziheng Li, Mozhen Cheng, Xiuling Chen, Aoxue Wang, Chao Wang, and Xiaoxuan Zhang. "Genome-Wide Identification of the U-Box E3 Ubiquitin Ligase Gene Family in Cabbage (Brassica oleracea var. capitata) and Its Expression Analysis in Response to Cold Stress and Pathogen Infection." Plants 12, no. 7 (March 24, 2023): 1437. http://dx.doi.org/10.3390/plants12071437.

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Plant U-box E3 ubiquitin ligases (PUBs) play an important role in growth, development, and stress responses in many species. However, the characteristics of U-box E3 ubiquitin ligase genes in cabbage (Brassica oleracea var. capitata) are still unclear. Here, we carry out the genome-wide analysis of U-box E3 ubiquitin ligase genes in cabbage and identify 65 Brassica oleracea var. capitata U-box E3 ubiquitin ligase (BoPUB) genes in the cabbage genome. Phylogenetic analysis indicates that all 65 BoPUB genes are grouped into six subfamilies, whose members are relatively conserved in the protein domain and exon-intron structure. Chromosomal localization and synteny analyses show that segmental and tandem duplication events contribute to the expansion of the U-box E3 ubiquitin ligase gene family in cabbage. Protein interaction prediction presents that heterodimerization may occur in BoPUB proteins. In silico promoter analysis and spatio-temporal expression profiling of BoPUB genes reveal their involvement in light response, phytohormone response, and growth and development. Furthermore, we find that BoPUB genes participate in the biosynthesis of cuticular wax and in response to cold stress and pathogenic attack. Our findings provide a deep insight into the U-box E3 ubiquitin ligase gene family in cabbage and lay a foundation for the further functional analysis of BoPUB genes in different biological processes.

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Tseng, Hui-Min, David Shum, Bhavneet Bhinder, Sindy Escobar, Nicholas J. Veomett, Alan E. Tomkinson, David Y. Gin, Hakim Djaballah, and David A. Scheinberg. "A High-Throughput Scintillation Proximity-Based Assay for Human DNA Ligase IV." ASSAY and Drug Development Technologies 10, no. 3 (June 2012): 235–49. http://dx.doi.org/10.1089/adt.2011.0404.

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McDermott, Jason E., John R. Cort, Ernesto S. Nakayasu, Jonathan N. Pruneda, Christopher Overall, and Joshua N. Adkins. "Prediction of bacterial E3 ubiquitin ligase effectors using reduced amino acid peptide fingerprinting." PeerJ 7 (June 7, 2019): e7055. http://dx.doi.org/10.7717/peerj.7055.

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Background Although pathogenic Gram-negative bacteria lack their own ubiquitination machinery, they have evolved or acquired virulence effectors that can manipulate the host ubiquitination process through structural and/or functional mimicry of host machinery. Many such effectors have been identified in a wide variety of bacterial pathogens that share little sequence similarity amongst themselves or with eukaryotic ubiquitin E3 ligases. Methods To allow identification of novel bacterial E3 ubiquitin ligase effectors from protein sequences we have developed a machine learning approach, the SVM-based Identification and Evaluation of Virulence Effector Ubiquitin ligases (SIEVE-Ub). We extend the string kernel approach used previously to sequence classification by introducing reduced amino acid (RED) alphabet encoding for protein sequences. Results We found that 14mer peptides with amino acids represented as simply either hydrophobic or hydrophilic provided the best models for discrimination of E3 ligases from other effector proteins with a receiver-operator characteristic area under the curve (AUC) of 0.90. When considering a subset of E3 ubiquitin ligase effectors that do not fall into known sequence based families we found that the AUC was 0.82, demonstrating the effectiveness of our method at identifying novel functional family members. Feature selection was used to identify a parsimonious set of 10 RED peptides that provided good discrimination, and these peptides were found to be located in functionally important regions of the proteins involved in E2 and host target protein binding. Our general approach enables construction of models based on other effector functions. We used SIEVE-Ub to predict nine potential novel E3 ligases from a large set of bacterial genomes. SIEVE-Ub is available for download at https://doi.org/10.6084/m9.figshare.7766984.v1 or https://github.com/biodataganache/SIEVE-Ub for the most current version.

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Vriend, Jerry, Thatchawan Thanasupawat, Namita Sinha, and Thomas Klonisch. "Ubiquitin Proteasome Gene Signatures in Ependymoma Molecular Subtypes." International Journal of Molecular Sciences 23, no. 20 (October 15, 2022): 12330. http://dx.doi.org/10.3390/ijms232012330.

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The ubiquitin proteasome system (UPS) is critically important for cellular homeostasis and affects virtually all key functions in normal and neoplastic cells. Currently, a comprehensive review of the role of the UPS in ependymoma (EPN) brain tumors is lacking but may provide valuable new information on cellular networks specific to different EPN subtypes and reveal future therapeutic targets. We have reviewed publicly available EPN gene transcription datasets encoding components of the UPS pathway. Reactome analysis of these data revealed genes and pathways that were able to distinguish different EPN subtypes with high significance. We identified differential transcription of several genes encoding ubiquitin E2 conjugases associated with EPN subtypes. The expression of the E2 conjugase genes UBE2C, UBE2S, and UBE2I was elevated in the ST_EPN_RELA subtype. The UBE2C and UBE2S enzymes are associated with the ubiquitin ligase anaphase promoting complex (APC/c), which regulates the degradation of substrates associated with cell cycle progression, whereas UBE2I is a Sumo-conjugating enzyme. Additionally, elevated in ST_EPN_RELA were genes for the E3 ligase and histone deacetylase HDAC4 and the F-box cullin ring ligase adaptor FBX031. Cluster analysis demonstrated several genes encoding E3 ligases and their substrate adaptors as EPN subtype specific genetic markers. The most significant Reactome Pathways associated with differentially expressed genes for E3 ligases and their adaptors included antigen presentation, neddylation, sumoylation, and the APC/c complex. Our analysis provides several UPS associated factors that may be attractive markers and future therapeutic targets for the subtype-specific treatment of EPN patients.

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Choi, Juyoung, Wonkyung Lee, Gynheung An, and Seong-Ryong Kim. "OsCBE1, a Substrate Receptor of Cullin4-Based E3 Ubiquitin Ligase, Functions as a Regulator of Abiotic Stress Response and Productivity in Rice." International Journal of Molecular Sciences 22, no. 5 (March 2, 2021): 2487. http://dx.doi.org/10.3390/ijms22052487.

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Ubiquitination is an important environmental stress response, and E3 ubiquitin ligases play a major role in the process. T-DNA insertion mutants of rice, Oscbe1-1, and Oscbe1-2, were identified through the screening of cold stress tolerance at seedling stage. Oscbe1 mutants showed a significantly higher cold stress tolerance in the fresh weight, chlorophyll content, and photosynthetic efficiency than wild type. Molecular prediction showed that OsCBE1 (Oryza sativa Cullin4-Based E3 ubiquitin ligase1) encoded a novel substrate receptor of Cullin4-based E3 ubiquitin ligase complex (C4E3). Whereas Oscbe1 mutants had fewer panicles and grains than wild type in the paddy field, the overexpression lines of OsCBE1 had more panicles and grains, suggesting that OsCBE1 is involved in the regulation of both abiotic stress response and development. Oscbe1 mutants also showed ABA hypersensitivity during seed germination, suggesting OsCBE1 function for the stress response via ABA signaling. In silico analysis of OsCBE1 activity predicted a CCCH-type transcription factor, OsC3H32, as a putative substrate. Co-IP (Co-immunoprecipitation) study showed that OsCBE1 interacts with OsDDB1, an expected binding component of OsCBE1 and OsC3H32. Additionally, expression of OsOLE16, OsOLE18, and OsBURP5 were negatively related with expression of OsCBE1. These results suggest that OsCBE1 functions as a regulator of the abiotic stress response via CCCH as a member of the C4E3.

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Sukumaran, Bindu, and Pradeep Bist. "Negative regulation of Nod2-mediated NF-κB activation (INM6P.417)." Journal of Immunology 192, no. 1_Supplement (May 1, 2014): 122.14. http://dx.doi.org/10.4049/jimmunol.192.supp.122.14.

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Abstract The cytoplasmic innate immune receptor NOD2 is required for innate immune defense against Gram-positive and Gram-negative intracellular bacterial pathogens. Dysregulation of NOD2 signaling is implicated in several inflammatory disorders (e.g., Crohn’s disease). However, a mechanistic global understanding of the negative regulation of this pathway needs further studies. Through a genome-scale RNA-interference screen, we identified 75 negative regulatory proteins of NOD1/2-mediated NF-κB activation. We characterized the role of several newly identified negative regulators in the regulation of NOD2-mediated NF-κB signaling, focusing on E3 ubiquitin ligases. Depletion or over expression of these ubiquitin ligases respectively enhanced or suppressed NOD2-mediated NF-κB activation, cytokine secretion and defense against intracellular bacteria. We identified and characterized a novel ubiquitin ligase that degrades Rip2, the adaptor of NOD2, via K48-linked ubiquitination, leading to negative feedback regulation. We also identified that ubiquitin ligase-mediated RIP2 degradation was essential for the tolerance in human immune cells observed following chronic NOD2 stimulation, providing a novel mechanism by which cells establish MDP tolerance. Our findings provide new insights into the mechanisms by which NOD2 pathway attenuated, and provide new directions to understand the role of NOD2 pathway in both infectious and inflammatory diseases.

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Tan, Xu, and Ning Zheng. "Hormone signaling through protein destruction: a lesson from plants." American Journal of Physiology-Endocrinology and Metabolism 296, no. 2 (February 2009): E223—E227. http://dx.doi.org/10.1152/ajpendo.90807.2008.

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Ubiquitin-dependent protein degradation has emerged as a major pathway regulating eukaryotic biology. By employing a variety of ubiquitin ligases to target specific cellular proteins, the ubiquitin-proteasome system controls physiological processes in a highly regulated fashion. Recent studies on a plant hormone auxin have unveiled a novel paradigm of signal transduction in which ubiquitin ligases function as hormone receptors. Perceived by the F-box protein subunit of the SCFTIR1 ubiquitin ligase, auxin directly promotes the recruitment of a family of transcriptional repressors for ubiquitination, thereby activating extensive transcriptional programs. Structural studies have revealed that auxin functions through a “molecular glue” mechanism to enhance protein-protein interactions with the assistance of another small molecule cofactor, inositol hexakisphosphate. Given the extensive repertoire of similar ubiquitin ligases in eukaryotic cells, this novel and widely adopted hormone-signaling mechanism in plants may also exist in other organisms.

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Yan, Yuqian, Jingwei Shao, Donglin Ding, Yunqian Pan, Phuc Tran, Wei Yan, Zhengyu Wang, Hong-yu Li, and Haojie Huang. "3-Aminophthalic acid, a new cereblon ligand for targeted protein degradation by O’PROTAC." Chemical Communications 58, no. 14 (2022): 2383–86. http://dx.doi.org/10.1039/d1cc06525d.

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In this study, we discovered 3-aminophthalic acid as a new ligand of cereblon (CRBN) E3 ubiquitin ligase and developed a phthalic acid-based O’PROTAC for ERG destruction, expanding the pool of ligands for development of PROTACs, especially O’PROTACs.

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Knop, Jan, Tim Lienemann, Haifa El-Kilani, Sven Falke, Catharina Krings, Maria Sindalovskaya, Johannes Bergler, Christian Betzel, and Stefan Hoth. "Structural Features of a Full-Length Ubiquitin Ligase Responsible for the Formation of Patches at the Plasma Membrane." International Journal of Molecular Sciences 22, no. 17 (August 31, 2021): 9455. http://dx.doi.org/10.3390/ijms22179455.

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Plant U-box armadillo repeat (PUB-ARM) ubiquitin (Ub) ligases have important functions in plant defense through the ubiquitination of target proteins. Defense against pathogens involves vesicle trafficking and the formation of extracellular vesicles. The PUB-ARM protein SENESCENCE ASSOCIATED UBIQUITIN E3 LIGASE1 (SAUL1) can form patches at the plasma membrane related to tethering multi-vesicular bodies (MVBs) to the plasma membrane. We uncovered the structure of a full-length plant ubiquitin ligase and the structural requirements of SAUL1, which are crucial for its function in patch formation. We resolved the structure of SAUL1 monomers by small-angle X-ray scattering (SAXS). The SAUL1 model showed that SAUL1 consists of two domains: a domain containing the N-terminal U-box and armadillo (ARM) repeats and the C-terminal ARM repeat domain, which includes a positively charged groove. We showed that all C-terminal ARM repeats are essential for patch formation and that this function requires arginine residue at position 736. By applying SAXS to polydisperse SAUL1 systems, the oligomerization of SAUL1 is detectable, with SAUL1 tetramers being the most prominent oligomers at higher concentrations. The oligomerization domain consists of the N-terminal U-box and some N-terminal ARM repeats. Deleting the U-box resulted in the promotion of the SAUL1 tethering function. Our findings indicate that structural changes in SAUL1 may be fundamental to its function in forming patches at the plasma membrane.

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Xu, Keheng, Nan Wu, Wenbo Yao, Xiaowei Li, Yonggang Zhou, and Haiyan Li. "The Biological Function and Roles in Phytohormone Signaling of the F-Box Protein in Plants." Agronomy 11, no. 11 (November 22, 2021): 2360. http://dx.doi.org/10.3390/agronomy11112360.

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The ubiquitin–proteasome pathway (UPP) is an important protein degradation pathway that can participate in the regulation of the physiological process of organisms by specifically removing abnormal peptides and degrading cell regulators. UPP mainly involves three enzymes, among which the E3 ubiquitin ligase function is central to UPP. E3 ubiquitin ligases can recruit substrate protein for ubiquitination, and they have various forms. Among them, the Skp1–Cul1–F-box (SCF) complex is the most representative member of the cullin RING ubiquitin ligases type in RING-domain E3 ligases, being mainly composed of Cullin 1, Skp1, Rbx1, and F-box proteins. The F-box protein is the key component for SCF to perform specific functions. The F-box protein is one of the largest protein families in plants, and its family members are involved in the regulation of many key physiological processes, such as growth and development of plants and the response to external stimuli. Herein, we briefly review the structure, classification, function, and hormone signaling pathways of F-box proteins.

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Anduro Corona, Iván, and Humberto Astiazaran García. "RNF8: ¿BLANCO TERAPÉUTICO POTENCIAL PARA TRATAR EL CÁNCER DE MAMA?" Biotecnia 20, no. 1 (January 15, 2018): 47–52. http://dx.doi.org/10.18633/biotecnia.v20i1.529.

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El cáncer de mama hereditario se ha asociado con alteraciones en el gen BRCA1, imposibilitando a la célula tumoral reparar las lesiones de doble cadena del ADN por recombinación homóloga. En la célula normal la RH es necesaria para el mantenimiento de la integridad del ADN. Sin embargo, en las células con BRCA1 disfuncional, el ADN es reparado por el sistema de unión de extremos no homólogos propenso a errores en la0reparación del ADN. Esta condición involucra a 53BP1, cuya función es esencial para el sistema UENH, favoreciendo la inestabilidad genómica y la tumorigénesis mamaria. RNF8 es una E3 ubiquitina ligasa que promueve el enlace de BRCA1 y 53BP1 ubicándolas en los sitios de ADN dañado. Se presentan una serie de alternativas con el objetivo de reconocer y promover la eliminación de RNF8. Estas aproximaciones presentan a RNF8 como blanco en estrategias farmacológicas para eliminar la inestabilidad genómica dependiente de 53BP1 y la resistencia farmacológica promovida por la inactivación de 53BP1 en células mamarias carentes de BRCA1.

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Nolte, Johannes Christoph, Marc Schürmann, Catherine-Louise Schepers, Elvira Vogel, Jan Hendrik Wübbeler, and Alexander Steinbüchel. "Novel Characteristics of Succinate Coenzyme A (Succinate-CoA) Ligases: Conversion of Malate to Malyl-CoA and CoA-Thioester Formation of Succinate AnaloguesIn Vitro." Applied and Environmental Microbiology 80, no. 1 (October 18, 2013): 166–76. http://dx.doi.org/10.1128/aem.03075-13.

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ABSTRACTThree succinate coenzyme A (succinate-CoA) ligases (SucCD) fromEscherichia coli,Advenella mimigardefordensisDPN7T, andAlcanivorax borkumensisSK2 were characterized regarding their substrate specificity concerning succinate analogues. Previous studies had suggested that SucCD enzymes might be promiscuous toward succinate analogues, such as itaconate and 3-sulfinopropionate (3SP). The latter is an intermediate of the degradation pathway of 3,3′-dithiodipropionate (DTDP), a precursor for the biotechnical production of polythioesters (PTEs) in bacteria. ThesucCDgenes were expressed inE. coliBL21(DE3)/pLysS. The SucCD enzymes ofE. coliandA. mimigardefordensisDPN7Twere purified in the native state using stepwise purification protocols, while SucCD fromA. borkumensisSK2 was equipped with a C-terminal hexahistidine tag at the SucD subunit. Besides the preference for the physiological substrates succinate, itaconate, ATP, and CoA, high enzyme activity was additionally determined for both enantiomeric forms of malate, amounting to 10 to 21% of the activity with succinate.Kmvalues ranged from 2.5 to 3.6 mM forl-malate and from 3.6 to 4.2 mM ford-malate for the SucCD enzymes investigated in this study. Asl-malate-CoA ligase is present in the serine cycle for assimilation of C1compounds in methylotrophs, structural comparison of these two enzymes as members of the same subsubclass suggested a strong resemblance of SucCD tol-malate-CoA ligase and gave rise to the speculation that malate-CoA ligases and succinate-CoA ligases have the same evolutionary origin. Although enzyme activities were very low for the additional substrates investigated, liquid chromatography/electrospray ionization-mass spectrometry analyses proved the ability of SucCD enzymes to form CoA-thioesters of adipate, glutarate, and fumarate. Since all SucCD enzymes were able to activate 3SP to 3SP-CoA, we consequently demonstrated that the activation of 3SP is not a unique characteristic of the SucCD fromA. mimigardefordensisDPN7T. The essential role ofsucCDin the activation of 3SPin vivowas proved by genetic complementation.

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Mallela, Abhishek, Maulik K. Nariya, and Eric J. Deeds. "Crosstalk and ultrasensitivity in protein degradation pathways." PLOS Computational Biology 16, no. 12 (December 28, 2020): e1008492. http://dx.doi.org/10.1371/journal.pcbi.1008492.

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Protein turnover is vital to cellular homeostasis. Many proteins are degraded efficiently only after they have been post-translationally “tagged” with a polyubiquitin chain. Ubiquitylation is a form of Post-Translational Modification (PTM): addition of a ubiquitin to the chain is catalyzed by E3 ligases, and removal of ubiquitin is catalyzed by a De-UBiquitylating enzyme (DUB). Nearly four decades ago, Goldbeter and Koshland discovered that reversible PTM cycles function like on-off switches when the substrates are at saturating concentrations. Although this finding has had profound implications for the understanding of switch-like behavior in biochemical networks, the general behavior of PTM cycles subject to synthesis and degradation has not been studied. Using a mathematical modeling approach, we found that simply introducing protein turnover to a standard modification cycle has profound effects, including significantly reducing the switch-like nature of the response. Our findings suggest that many classic results on PTM cycles may not hold in vivo where protein turnover is ubiquitous. We also found that proteins sharing an E3 ligase can have closely related changes in their expression levels. These results imply that it may be difficult to interpret experimental results obtained from either overexpressing or knocking down protein levels, since changes in protein expression can be coupled via E3 ligase crosstalk. Understanding crosstalk and competition for E3 ligases will be key in ultimately developing a global picture of protein homeostasis.

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Woo, Jennifer L., and Arnold J. Berk. "Adenovirus Ubiquitin-Protein Ligase Stimulates Viral Late mRNA NuclearExport." Journal of Virology 81, no. 2 (January 15, 2007): 575–87. http://dx.doi.org/10.1128/jvi.01725-06.

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ABSTRACT Theadenovirus type 5 (Ad5) E1B-55K and E4orf6 proteins are required together to stimulate viral late nuclear mRNA export to the cytoplasm and to restrict host cell nuclear mRNA export during the late phase of infection. Previous studies have shown that these two viral proteins interact with the cellular proteins elongins B and C, cullin 5, RBX1, and additional cellular proteins to form an E3 ubiquitin-protein ligase that polyubiquitinates p53 and probably one or more subunits of the MRE11-RAD50-NBS1 (MRN) complex, directing their proteasomal degradation. The MRN complex is required for cellular DNA double-strand break repair and induction of the DNA damage response by adenovirus infection. To determine if the ability of E1B-55K and E4orf6 to stimulate viral late mRNA nuclear export requires the ubiquitin-protein ligase activity of this viral ubiquitin-protein ligase complex, we designed and expressed a dominant-negative mutant form of cullin 5 in HeLa cells before infection with wild-type Ad5 or the E1B-55K null mutant dl1520. The dominant-negative cullin 5 protein stabilized p53 and the MRN complex, indicating that it inhibited the viral ubiquitin-protein ligase but had no effect on viral early mRNA synthesis, early protein synthesis, or viral DNA replication. However, expression of the dominant-negative cullin 5 protein caused a decrease in viral late protein synthesis and viral nuclear mRNA export similar to the phenotype produced by mutations in E1B-55K. We conclude that the stimulation of adenovirus late mRNA nuclear export by E1B-55K and E4orf6 results from the ubiquitin-protein ligase activity of the adenovirus ubiquitin-protein ligase complex.

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Heras, Gabriel, Arvind Venkat Namuduri, Leonardo Traini, Ganna Shevchenko, Alexander Falk, Sara Bergström Lind, Mi Jia, Geng Tian, and Stefano Gastaldello. "Muscle RING-finger protein-1 (MuRF1) functions and cellular localization are regulated by SUMO1 post-translational modification." Journal of Molecular Cell Biology 11, no. 5 (June 4, 2018): 356–70. http://dx.doi.org/10.1093/jmcb/mjy036.

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Abstract The muscle RING-finger protein-1 (MuRF1) is an E3 ubiquitin ligase expressed in skeletal and cardiac muscle tissues and it plays important roles in muscle remodeling. Upregulation of MuRF1 gene transcription participates in skeletal muscle atrophy, on contrary downregulation of protein expression leads to cardiac hypertrophy. MuRF1 gene point mutations have been found to generate protein aggregate myopathies defined as muscle disorder characterized by protein accumulation in muscle fibers. We have discovered that MuRF1 turned out to be also a target for a new post-translational modification arbitrated by conjugation of SUMO1 and it is mediated by the SUMO ligases E2 UBC9 and the E3 PIASγ/4. SUMOylation takes place at lysine 238 localized at the second coiled-coil protein domain that is required for efficient substrate interaction for polyubiquitination. We provided evidence that SUMOylation is essential for MuRF1 nuclear translocation and its mitochondria accumulation is enhanced in hyperglycemic conditions delivering a stabilization of the overall SUMOylated proteins in cultured myocytes. Thus, our findings add this SUMO1 post-translational modification as a new concept to understand muscle disorders related to the defect in MuRF1 activity.

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Cassaday, Jason, Tarak Shah, Justin Murray, Gregory T. O'Donnell, Oleg Kornienko, Berta Strulovici, Marc Ferrer, and Paul Zuck. "Miniaturization and Automation of an Ubiquitin Ligase Cascade Enzyme-Linked Immunosorbent Assay in 1,536-Well Format." ASSAY and Drug Development Technologies 5, no. 4 (August 2007): 493–500. http://dx.doi.org/10.1089/adt.2007.076.

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St-Pierre, Patrick, Euan Shaw, Samuel Jacques, Paul A. Dalgarno, Cibran Perez-Gonzalez, Frédéric Picard-Jean, J. Carlos Penedo, and Daniel A. Lafontaine. "A structural intermediate pre-organizes the add adenine riboswitch for ligand recognition." Nucleic Acids Research 49, no. 10 (May 8, 2021): 5891–904. http://dx.doi.org/10.1093/nar/gkab307.

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Abstract Riboswitches are RNA sequences that regulate gene expression by undergoing structural changes upon the specific binding of cellular metabolites. Crystal structures of purine-sensing riboswitches have revealed an intricate network of interactions surrounding the ligand in the bound complex. The mechanistic details about how the aptamer folding pathway is involved in the formation of the metabolite binding site have been previously shown to be highly important for the riboswitch regulatory activity. Here, a combination of single-molecule FRET and SHAPE assays have been used to characterize the folding pathway of the adenine riboswitch from Vibrio vulnificus. Experimental evidences suggest a folding process characterized by the presence of a structural intermediate involved in ligand recognition. This intermediate state acts as an open conformation to ensure ligand accessibility to the aptamer and folds into a structure nearly identical to the ligand-bound complex through a series of structural changes. This study demonstrates that the add riboswitch relies on the folding of a structural intermediate that pre-organizes the aptamer global structure and the ligand binding site to allow efficient metabolite sensing and riboswitch genetic regulation.

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Lama, Rati, Samuel L. Galster, Chao Xu, Luke W. Davison, Sherry R. Chemler, and Xinjiang Wang. "Dual Targeting of MDM4 and FTH1 by MMRi71 for Induced Protein Degradation and p53-Independent Apoptosis in Leukemia Cells." Molecules 27, no. 22 (November 8, 2022): 7665. http://dx.doi.org/10.3390/molecules27227665.

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MDM2 and MDM4 are cancer drug targets validated in multiple models for p53-based cancer therapies. The RING domains of MDM2 and non-p53-binder MDM2 splice isoforms form RING domain heterodimer polyubiquitin E3 ligases with MDM4, which regulate p53 stability in vivo and promote tumorigenesis independent of p53. Despite the importance of the MDM2 RING domain in p53 regulation and cancer development, small molecule inhibitors targeting the E3 ligase activity of MDM2-MDM4 are poorly explored. Here, we describe the synthesis and characterization of quinolinol derivatives for the identification of analogs that are capable of targeting the MDM2-MDM4 heterodimer E3 ligase and inducing apoptosis in cells. The structure-activity-relationship (SAR) study identified structural moieties critical for the inhibitory effects toward MDM2-MDM4 E3 ligase, the targeted degradation of MDM4 and FTH1 in cells, and anti-proliferation activity. Lead optimization led to the development of compound MMRi71 with improved activity. In addition to accumulating p53 proteins in wt-p53 bearing cancer cells as expected of any MDM2 inhibitors, MMRi71 effectively kills p53-null leukemia cells, an activity that conventional MDM2-p53 disrupting inhibitors lack. This study provides a prototype structure for developing MDM4/FTH1 dual-targeting inhibitors as potential cancer therapeutics.

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Goettig, Peter. "Reversed Proteolysis—Proteases as Peptide Ligases." Catalysts 11, no. 1 (December 30, 2020): 33. http://dx.doi.org/10.3390/catal11010033.

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Historically, ligase activity by proteases was theoretically derived due to their catalyst nature, and it was experimentally observed as early as around 1900. Initially, the digestive proteases, such as pepsin, chymotrypsin, and trypsin were employed to perform in vitro syntheses of small peptides. Protease-catalyzed ligation is more efficient than peptide bond hydrolysis in organic solvents, representing control of the thermodynamic equilibrium. Peptide esters readily form acyl intermediates with serine and cysteine proteases, followed by peptide bond synthesis at the N-terminus of another residue. This type of reaction is under kinetic control, favoring aminolysis over hydrolysis. Although only a few natural peptide ligases are known, such as ubiquitin ligases, sortases, and legumains, the principle of proteases as general catalysts could be adapted to engineer some proteases accordingly. In particular, the serine proteases subtilisin and trypsin were converted to efficient ligases, which are known as subtiligase and trypsiligase. Together with sortases and legumains, they turned out to be very useful in linking peptides and proteins with a great variety of molecules, including biomarkers, sugars or building blocks with non-natural amino acids. Thus, these engineered enzymes are a promising branch for academic research and for pharmaceutical progress.

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Kim, Jae Ho, Moon Seok Kim, Dae Yeon Kim, Joseph Noble Amoah, and Yong Weon Seo. "Molecular Characterization of U-box E3 Ubiquitin Ligases (TaPUB2 and TaPUB3) Involved in the Positive Regulation of Drought Stress Response in Arabidopsis." International Journal of Molecular Sciences 22, no. 24 (December 20, 2021): 13658. http://dx.doi.org/10.3390/ijms222413658.

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Plant U-box E3 ubiquitin ligase (PUB) is involved in various environmental stress conditions. However, the molecular mechanism of U-box proteins in response to abiotic stress in wheat remains unknown. In this study, two U-box E3 ligase genes (TaPUB2 and TaPUB3), which are highly expressed in response to adverse abiotic stresses, were isolated from common wheat, and their cellular functions were characterized under drought stress. Transient expression assay revealed that TaPUB2 was localized in the cytoplasm and Golgi apparatus, whereas TaPUB3 was expressed only in the Golgi apparatus in wheat protoplasts. Additionally, TaPUB2 and TaPUB3 underwent self-ubiquitination. Moreover, TaPUB2/TaPUB3 heterodimer was identified in yeast and the cytoplasm of wheat protoplasts using a pull-down assay and bimolecular fluorescence complementation analysis. Heterogeneous overexpression of TaPUB2 and TaPUB3 conferred tolerance to drought stress. Taken together, these results implied that the heterodimeric form of U-box E3 ubiquitin ligases (TaPUB2/TaPUB3) responded to abiotic stress and roles as a positive regulator of drought stress tolerance.

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Li, Jun-Mei, Mei-Yan Ye, Chaofeng Wang, Xiao-Han Ma, Ni-Ni Wu, Chen-Li Zhong, Yanjun Zhang, et al. "Soybean GmSAUL1, a Bona Fide U-Box E3 Ligase, Negatively Regulates Immunity Likely through Repressing the Activation of GmMPK3." International Journal of Molecular Sciences 24, no. 7 (March 25, 2023): 6240. http://dx.doi.org/10.3390/ijms24076240.

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E3 ubiquitin ligases play important roles in plant immunity, but their role in soybean has not been investigated previously. Here, we used Bean pod mottle virus (BPMV)-mediated virus-induced gene silencing (VIGS) to investigate the function of GmSAUL1 (Senescence-Associated E3 Ubiquitin Ligase 1) homologs in soybean. When two closely related SAUL1 homologs were silenced simultaneously, the soybean plants displayed autoimmune phenotypes, which were significantly alleviated by high temperature, suggesting that GmSAUL1a/1b might be guarded by an R protein. Interestingly, silencing GmSAUL1a/1b resulted in the decreased activation of GmMPK6, but increased activation of GmMPK3 in response to flg22, suggesting that the activation of GmMPK3 is most likely responsible for the activated immunity observed in the GmSAUL1a/1b-silenced plants. Furthermore, we provided evidence that GmSAUL1a is a bona fide E3 ligase. Collectively, our results indicated that GmSAUL1 plays a negative role in regulating cell death and immunity in soybean.

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Castillo-Villanueva, Elizabeth, Grisel Ballesteros, Melanie Schmid, Paloma Hidalgo, Sabrina Schreiner, Thomas Dobner, and Ramon A. Gonzalez. "The Mre11 Cellular Protein Is Modified by Conjugation of Both SUMO-1 and SUMO-2/3 during Adenovirus Infection." ISRN Virology 2014 (April 7, 2014): 1–14. http://dx.doi.org/10.1155/2014/989160.

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The adenovirus type 5 (Ad5) E1B 55 kDa and E4 Orf6 proteins assemble a Cullin 5-E3 ubiquitin (Ub) ligase that targets, among other cellular proteins, p53 and the Mre11-Rad50-Nbs1 (MRN) complex for degradation. The latter is also inhibited by the E4 Orf3 protein, which promotes the recruitment of Mre11 into specific nuclear sites to promote viral DNA replication. The activities associated with the E1B 55 kDa and E4 Orf6 viral proteins depend mostly on the assembly of this E3-Ub ligase. However, E1B 55 kDa can also function as an E3-SUMO ligase, suggesting not only that regulation of cellular proteins by these viral early proteins may depend on polyubiquitination and proteasomal degradation but also that SUMOylation of target proteins may play a key role in their activities. Since Mre11 is a target of both the E1B/E4 Orf6 complex and E4 Orf3, we decided to determine whether Mre11 displayed similar properties to those of other cellular targets, in Ad5-infected cells. We have found that during Ad5-infection, Mre11 is modified by SUMO-1 and SUMO-2/3 conjugation. Unexpectedly, SUMOylation of Mre11 is not exclusively dependent on E1B 55 kDa, E4 Orf6, or E4 Orf3, rather it seems to be influenced by a molecular interplay that involves each of these viral early proteins.

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Tracz, Michał, Ireneusz Górniak, Andrzej Szczepaniak, and Wojciech Białek. "E3 Ubiquitin Ligase SPL2 Is a Lanthanide-Binding Protein." International Journal of Molecular Sciences 22, no. 11 (May 27, 2021): 5712. http://dx.doi.org/10.3390/ijms22115712.

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The SPL2 protein is an E3 ubiquitin ligase of unknown function. It is one of only three types of E3 ligases found in the outer membrane of plant chloroplasts. In this study, we show that the cytosolic fragment of SPL2 binds lanthanide ions, as evidenced by fluorescence measurements and circular dichroism spectroscopy. We also report that SPL2 undergoes conformational changes upon binding of both Ca2+ and La3+, as evidenced by its partial unfolding. However, these structural rearrangements do not interfere with SPL2 enzymatic activity, as the protein retains its ability to auto-ubiquitinate in vitro. The possible applications of lanthanide-based probes to identify protein interactions in vivo are also discussed. Taken together, the results of this study reveal that the SPL2 protein contains a lanthanide-binding site, showing for the first time that at least some E3 ubiquitin ligases are also capable of binding lanthanide ions.

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Baek, Suk-Hwan, Bin Huang, and Hyeun Wook Chang. "RNF144b is a negative regulator in TLR2-mediated NF-kB activation." Journal of Immunology 196, no. 1_Supplement (May 1, 2016): 132.4. http://dx.doi.org/10.4049/jimmunol.196.supp.132.4.

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Abstract NF-κB regulates the expression of a various genes involved in diverse cellular processes including inflammation and immunity. Activation of NF-κB requires ubiquitination, a highly conserved and versatile modification that can regulate cell signaling through both proteasome dependent and independent mechanisms. Ubiquitination process is a representative of post-translational modification involved in NF-κB activation of TNFR and TLR signaling. We demonstrated that RNF144b, an E3 ubiquitin ligase, is important factor for the NF-κB regulation in macrophages upon stimulation with TNFa or TLR2 ligand. Treatment of TNFa or TLR2 ligand (Pam3CSK4) decreased the expression of RNF144b mRNA on macrophages. TLR2 ligand has a stronger effect than TNFa in the expression of RNF144b mRNA. Knock-down of RNF144b potentiated activation of NF-kB. And according to the increase of NF-κB activity, the formation of foam cells and the production of cytokines (IL-1β, IL-6, CCL2 and CCL3) following TLR2 stimulation increased. On the contrary, overexpression of RNF144b decreased NF-κB activity and cytokines production in the TLR2-stimuated macrophages. RNF144b binds and ubiquitinates TRAF6 and these effects are dependent of its E3 ligase activity. Mutation of Cys201 to Ala abolishes activity of RNF144b to activate TRAF6 ubiquitination and also partially attenuates NF-κB activity. Taken together, these results suggest that RNF144b stimulates NF-κB activity via TRAF6 ubiquitination and then produces inflammatory cytokines and chemokines, and eventually controls foam cell formation in the TLR2-mediated macrophages.

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Kats, Ilia, Christian Reinbold, Marc Kschonsak, Anton Khmelinskii, Laura Armbruster, Thomas Ruppert, and Michael Knop. "Up-regulation of ubiquitin–proteasome activity upon loss of NatA-dependent N-terminal acetylation." Life Science Alliance 5, no. 2 (November 11, 2021): e202000730. http://dx.doi.org/10.26508/lsa.202000730.

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N-terminal acetylation is a prominent protein modification, and inactivation of N-terminal acetyltransferases (NATs) cause protein homeostasis stress. Using multiplexed protein stability profiling with linear ubiquitin fusions as reporters for the activity of the ubiquitin proteasome system, we observed increased ubiquitin proteasome system activity in NatA, but not NatB or NatC mutants. We find several mechanisms contributing to this behavior. First, NatA-mediated acetylation of the N-terminal ubiquitin–independent degron regulates the abundance of Rpn4, the master regulator of the expression of proteasomal genes. Second, the abundance of several E3 ligases involved in degradation of UFD substrates is increased in cells lacking NatA. Finally, we identify the E3 ligase Tom1 as a novel chain-elongating enzyme (E4) involved in the degradation of linear ubiquitin fusions via the formation of branched K11, K29, and K48 ubiquitin chains, independently of the known E4 ligases involved in UFD, leading to enhanced ubiquitination of the UFD substrates.

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Yi, So Young, Myungjin Lee, Suk-Yoon Kwon, Woo Taek Kim, Yong Pyo Lim, and Si-Yong Kang. "RING-Type E3 Ubiquitin Ligases AtRDUF1 and AtRDUF2 Positively Regulate the Expression of PR1 Gene and Pattern-Triggered Immunity." International Journal of Molecular Sciences 23, no. 23 (November 22, 2022): 14525. http://dx.doi.org/10.3390/ijms232314525.

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The importance of E3 ubiquitin ligases from different families for plant immune signaling has been confirmed. Plant RING-type E3 ubiquitin ligases are members of the E3 ligase superfamily and have been shown to play positive or negative roles during the regulation of various steps of plant immunity. Here, we present Arabidopsis RING-type E3 ubiquitin ligases AtRDUF1 and AtRDUF2 which act as positive regulators of flg22- and SA-mediated defense signaling. Expression of AtRDUF1 and AtRDUF2 is induced by pathogen-associated molecular patterns (PAMPs) and pathogens. The atrduf1 and atrduf2 mutants displayed weakened responses when triggered by PAMPs. Immune responses, including oxidative burst, mitogen-activated protein kinase (MAPK) activity, and transcriptional activation of marker genes, were attenuated in the atrduf1 and atrduf2 mutants. The suppressed activation of PTI responses also resulted in enhanced susceptibility to bacterial pathogens. Interestingly, atrduf1 and atrduf2 mutants showed defects in SA-mediated or pathogen-mediated PR1 expression; however, avirulent Pseudomonas syringae pv. tomato DC3000-induced cell death was unaffected. Our findings suggest that AtRDUF1 and AtRDUF2 are not just PTI-positive regulators but are also involved in SA-mediated PR1 gene expression, which is important for resistance to P. syringae.

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Sheppard, Hilary M., Janet C. Harries, Sagair Hussain, Charlotte Bevan, and David M. Heery. "Analysis of the Steroid Receptor Coactivator 1 (SRC1)-CREB Binding Protein Interaction Interface and Its Importance for the Function of SRC1." Molecular and Cellular Biology 21, no. 1 (January 1, 2001): 39–50. http://dx.doi.org/10.1128/mcb.21.1.39-50.2001.

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ABSTRACT The transcriptional activity of nuclear receptors is mediated by coactivator proteins, including steroid receptor coactivator 1 (SRC1) and its homologues and the general coactivators CREB binding protein (CBP) and p300. SRC1 contains an activation domain (AD1) which functions via recruitment of CBP and and p300. In this study, we have used yeast two-hybrid and in vitro interaction-peptide inhibition experiments to map the AD1 domain of SRC1 to a 35-residue sequence potentially containing two α-helices. We also define a 72-amino-acid sequence in CBP necessary for SRC1 binding, designated the SRC1 interaction domain (SID). We show that in contrast to SRC1, direct binding of CBP to the estrogen receptor is weak, suggesting that SRC1 functions primarily as an adaptor to recruit CBP and p300. In support of this, we show that the ability of SRC1 to enhance ligand-dependent nuclear receptor activity in transiently transfected cells is dependent upon the integrity of the AD1 region. In contrast, the putative histone acetyltransferase domain, the Per-Arnt-Sim basic helix-loop-helix domain, the glutamine-rich domain, and AD2 can each be removed without loss of ligand-induced activity. Remarkably, a construct corresponding to residues 631 to 970, which contains only the LXXLL motifs and the AD1 region of SRC1, retained strong coactivator activity in our assays.

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Nakotte, Tom, Hongmei Luo, and Jeff Pietryga. "Carrier Density Modulation in PbSe Quantum Dot Films via In-Solution Ligand Exchange." MRS Advances 5, no. 40-41 (2020): 2091–99. http://dx.doi.org/10.1557/adv.2020.254.

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AbstractIn-solution ligand exchange of PbSe QDs is used to examine the effect of capping ligand on the carrier density of PbSe QD films. Results show that carrier density can be modulated by a factor of 5 by choice of ligand without any additional post deposition treatments. Proper fabrication and measurement conditions for calculating carrier densities from C-V measurements using a sandwich structure on P-doped Si/SiO2/Al2O3/QD/Au structure capacitance devices are outlined. Combining carrier density results with field-effect-transistor measurements, promising ligands which display lower carrier densities without having a significant drop off in carrier mobility are identified as candidates for photodetection purposes.

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Kołaczkowski, M., P. Mierzejewski, P. Bieńkowski, A. Wesołowska, and A. Newman-Tancredi. "ADN-1184 a monoaminergic ligand with 5-HT6/7receptor antagonist activity: pharmacological profile and potential therapeutic utility." British Journal of Pharmacology 171, no. 4 (January 24, 2014): 973–84. http://dx.doi.org/10.1111/bph.12509.

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El-Saafin, Farrah, Didier Devys, Steven A. Johnsen, Stéphane D. Vincent, and László Tora. "SAGA-Dependent Histone H2Bub1 Deubiquitination Is Essential for Cellular Ubiquitin Balance during Embryonic Development." International Journal of Molecular Sciences 23, no. 13 (July 5, 2022): 7459. http://dx.doi.org/10.3390/ijms23137459.

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Ubiquitin (ub) is a small, highly conserved protein widely expressed in eukaryotic cells. Ubiquitination is a post-translational modification catalyzed by enzymes that activate, conjugate, and ligate ub to proteins. Substrates can be modified either by addition of a single ubiquitin molecule (monoubiquitination), or by conjugation of several ubs (polyubiquitination). Monoubiquitination acts as a signaling mark to control diverse biological processes. The cellular and spatial distribution of ub is determined by the opposing activities of ub ligase enzymes, and deubiquitinases (DUBs), which remove ub from proteins to generate free ub. In mammalian cells, 1–2% of total histone H2B is monoubiquitinated. The SAGA (Spt Ada Gcn5 Acetyl-transferase) is a transcriptional coactivator and its DUB module removes ub from H2Bub1. The mammalian SAGA DUB module has four subunits, ATXN7, ATXN7L3, USP22, and ENY2. Atxn7l3−/− mouse embryos, lacking DUB activity, have a five-fold increase in H2Bub1 retention, and die at mid-gestation. Interestingly, embryos lacking the ub encoding gene, Ubc, have a similar phenotype. Here we provide a current overview of data suggesting that H2Bub1 retention on the chromatin in Atxn7l3−/− embryos may lead to an imbalance in free ub distribution. Thus, we speculate that ATXN7L3-containing DUBs impact the free cellular ub pool during development.

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Journal articles on the topic 'ADN Ligase'

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