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Journal articles on the topic "Rapid resonance assignment of proteins"
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Fredriksson, Jonas, Wolfgang Bermel, and Martin Billeter. "Complete protein assignment from sets of spectra recorded overnight." Journal of Biomolecular NMR 73, no. 1-2 (February 15, 2019): 59–70. http://dx.doi.org/10.1007/s10858-019-00226-8.
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Abstract A flexible and scalable approach for protein NMR is introduced that builds on rapid data collection via projection spectroscopy and analysis of the spectral input data via joint decomposition. Input data may originate from various types of spectra, depending on the ultimate goal: these may result from experiments based on triple-resonance pulse sequences, or on TOCSY or NOESY sequences, or mixtures thereof. Flexible refers to the free choice of spectra for the joint decompositions depending on the purpose: assignments, structure, dynamics, interactions. Scalable means that the approach is open to the addition of similar or different experiments, e.g. larger proteins may require a wider selection of triple-resonance based experiments. Central to the proposed approach is the mutual support among the different spectra during the spectral analysis: for example, sparser triple-resonance spectra may help decomposing (separating) spin systems in a TOCSY or identifying unique NOEs. In the example presented, backbone plus side chain assignments of ubiquitin were obtained from the combination of either two or three of the following projection experiments: a 4D HCCCONH, a 4D HNCACO and a 3D HNCACB. In all cases, TOCSY data (4D HCCCONH) proved crucial not only for the side chain assignments, but also for the sequential assignment. Even when total recording time was reduced to about 10 h, nearly complete assignments were obtained, with very few missing assignments and even fewer differences to a reference.
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Rout, Ashok K., Ravi P. Barnwal, Geetika Agarwal, and Kandala V. R. Chary. "Root-mean-square-deviation-based rapid backbone resonance assignments in proteins." Magnetic Resonance in Chemistry 48, no. 10 (August 27, 2010): 793–97. http://dx.doi.org/10.1002/mrc.2664.
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Sukumaran, Sujeesh, Shahid A. Malik, Shankararama Sharma R., Kousik Chandra, and Hanudatta S. Atreya. "Rapid NMR assignments of intrinsically disordered proteins using two-dimensional13C-detection based experiments." Chemical Communications 55, no. 54 (2019): 7820–23. http://dx.doi.org/10.1039/c9cc03530c.
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Chatterjee, Amarnath, Neel S. Bhavesh, Sanjay C. Panchal, and Ramakrishna V. Hosur. "A novel protocol based on HN(C)N for rapid resonance assignment in (15N, 13C) labeled proteins: implications to structural genomics." Biochemical and Biophysical Research Communications 293, no. 1 (April 2002): 427–32. http://dx.doi.org/10.1016/s0006-291x(02)00240-1.
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Kostic, Milka, Susan Sondej Pochapsky, and Thomas C. Pochapsky. "Rapid Recycle13C‘,15N and13C,13C‘ Heteronuclear and Homonuclear Multiple Quantum Coherence Detection for Resonance Assignments in Paramagnetic Proteins: Example of Ni2+-Containing Acireductone Dioxygenase." Journal of the American Chemical Society 124, no. 31 (August 2002): 9054–55. http://dx.doi.org/10.1021/ja0268480.
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Vendrell, J., F. X. Avilés, M. Vilanova, C. H. Turner, and C. Crane-Robinson. "1H-n.m.r. studies of the isolated activation segment from pig procarboxypeptidase A." Biochemical Journal 267, no. 1 (April 1, 1990): 213–20. http://dx.doi.org/10.1042/bj2670213.
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The isolated activation segment (asA) from pig pancreatic procarboxypeptidase A was studied by 1H-n.m.r. spectroscopy over a wide range of solution conditions. Isolated asA shows many characteristics of compactly folded globular proteins, such as the observation of perturbed positions for resonances from methyl groups, alpha-carbon atoms, histidine residues and the tyrosine residue. The single tyrosine residue (Tyr-70) exhibits a very high pKa, and both histidine and tyrosine residues show slow chemical modification (deuteration and iodination). In contrast, asA shows rapid NH exchange. Analysis of the spectra by pH titration and nuclear Overhauser effects revealed several residue interactions. Quantitative analysis of deuterium and tritium exchange allowed the assignment of the histidine C-2-H resonances to their respective residues in the sequence. His-66, the closest to the sites of proteolytic attack in the proenzyme, is shown to be the most accessible to solvent in procarboxypeptidase A. It was also shown that asA is thermally very stable [‘melting’ temperature (Tm) 88 degrees C] and requires a high urea concentration for denaturation (6.25 M, at pH 7.5). Evidence is presented for some degree of conformational flexibility in the premelting range, a feature that could be ascribed to the preponderance of helical secondary structure and to the lack of disulphide bridges. The free solution structure of asA is probably unchanged when it binds to carboxypeptidase A.
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Kumar, Dinesh, and Ramakrishna V. Hosur. "hNCOcanH pulse sequence and a robust protocol for rapid and unambiguous assignment of backbone (1 HN , 15 N and 13 C′) resonances in 15 N/13 C-labeled proteins." Magnetic Resonance in Chemistry 49, no. 9 (August 5, 2011): 575–83. http://dx.doi.org/10.1002/mrc.2787.
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Fiorito, Francesco, Sebastian Hiller, Gerhard Wider, and Kurt Wüthrich. "Automated Resonance Assignment of Proteins: 6 DAPSY-NMR." Journal of Biomolecular NMR 35, no. 1 (May 2006): 27–37. http://dx.doi.org/10.1007/s10858-006-0030-x.
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Higman, Victoria A. "Solid-state MAS NMR resonance assignment methods for proteins." Progress in Nuclear Magnetic Resonance Spectroscopy 106-107 (June 2018): 37–65. http://dx.doi.org/10.1016/j.pnmrs.2018.04.002.
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Crippen, Gordon M., Aikaterini Rousaki, Matthew Revington, Yongbo Zhang, and Erik R. P. Zuiderweg. "SAGA: rapid automatic mainchain NMR assignment for large proteins." Journal of Biomolecular NMR 46, no. 4 (March 16, 2010): 281–98. http://dx.doi.org/10.1007/s10858-010-9403-2.
Full textDissertations / Theses on the topic "Rapid resonance assignment of proteins"
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Jung, Young-Sang. "Rapid determination of protein structures in solution using NMR dipolar couplings." Doctoral thesis, [S.l.] : [s.n.], 2005. http://webdoc.sub.gwdg.de/diss/2005/jung/jung.pdf.
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Li, Kuo-Bin. "Development of computer-assisted methods for the resonance assignment of heteronuclear 3D NMR spectra of proteins." Thesis, McGill University, 1996. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=40381.
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An automated sequential assignment protocol for proteins is presented using heteronuclear 3D NMR. For the observed amino acid spin systems, the protocol includes an algorithm to determine their amino acid types. For the detected polypeptides, the protocol includes another algorithm to sequentially map them to the primary sequence. The former algorithm measures the similarity between the detected spin systems and the 20 standard amino acid patterns. Both chemical shift and topological likeness are considered. Knowing the amino acid types, the mapping algorithm assigns the detected polypeptides to proper positions within the protein primary sequence. The assignment protocol can be applied to spin systems generated from many different approaches. To demonstrate the assignment protocol, a few computer algorithms were designed to deduce the backbone and side-chain spin systems of proteins using heteronuclear 3D NMR. Magnetization transfer through peptide bonds can be observed in triple resonance 3D NMR. To automate the backbone assignment using the through-bond correlations, a generic algorithm is proposed. This algorithm searches and merges cross peaks among all available NMR spectra. Individual spin systems can be extracted and linked to create polypeptide chains based on the observed interresidue correlations. The algorithm is not restricted to any particular type of experiment. It is shown to be applicable to two sets of NMR spectra: the five experiment set of 3D HNCO, HNCA, HN(CO)CA, HCACO, $ sp{15}$N TOCSY-HMQC and the one-experiment set of 3D CBCANH. For the side chain assignment, an automated approach using a constrained partitioning algorithm has been developed to extract side chain spin systems of proteins by analyzing the 3D HCCH-COSY/TOCSY spectra. The extracted amino acid spin systems show the chemical shifts of the component nuclear spins as well as the connectivities between these spins. A 90-residue protein, the N-domain of chicken skeletal troponin-C
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Helgstrand, Magnus. "Structure determination of ribosomal proteins and development of new methods in biomolecular NMR." Doctoral thesis, Stockholm : Tekniska högsk, 2001. http://urn.kb.se/resolve?urn=urn:nbn:se:kth:diva-3149.
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Habenstein, Birgit. "Structural insights into fibrillar proteins from solid-state NMR spectroscopy." Thesis, Lyon 1, 2011. http://www.theses.fr/2011LYO10212.
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La RMN à l’état solide est une méthode de choix pour l’étude des protéines insolubles et des complexes protéiques de haut poids moléculaire. L’insolubilité intrinsèque des protéines fibrillaires, ainsi que leur architecture complexe, rendent difficile leur caractérisation structurale par la cristallographie et par la RMN en solution. La RMN à l‘état solide n’est pas limitée par le poids moléculaire et constitue donc un outil puissant pour l’étude des protéines fibrillaires. L’attribution des résonances RMN est le prérequis pour obtenir des informations structurales à résolution atomique. La première partie de ce travail de thèse décrit le développement de méthodes en RMN à l’état solide pour l’attribution des résonances. Nous avons appliqué ces méthodes afin d’attribuer le domaine C-terminal du prion Ure2 (33 kDa), qui est à ce jour la plus grande protéine attribuée par RMN à l’état solide. Nos résultats fournissent les bases pour l’étude de protéines à haut poids moléculaire à l’échelle atomique. Ceci est démontré dans la seconde partie de ce travail de thèse avec les premières études RMN à l’état solide des fibrilles des prions Ure2 et Sup35. Nous avons caractérisé la structure de ces prions pour les fibrilles entières ainsi que pour les domaines isolés. La troisième fibrille étudiée est l’α- synuclein, fibrille associée à la maladie de Parkinson, pour laquelle nous présentons l’attribution des résonances RMN ainsi que la structure secondaire d’un nouveau polymorphe. Les études présentées ici fournissent de nouvelles clés pour comprendre la diversité des architectures de fibrilles, en considérant les fibrilles comme entités individuelles d’un point de vue structuralSolid-state NMR is the method of choice for studies on insoluble proteins and other high molecular weight protein complexes. The inherent insolubility of fibrillar proteins, as well as their complex architecture, makes the application of x-ray crystallography and solution state NMR difficult. Solid-state NMR is not limited by the molecular weight or by the absence of long-range structural order, and is thus a powerful tool for the 3D structural investigation of fibrillar proteins. The assignment of the NMR resonances is a prerequisite to obtain structural information at atomic level. The first part of this thesis describes the development of solid-state NMR methods to assign the resonances in large proteins. We apply these methods to assign the 33 kDa C-terminal domain of the Ure2p prion which is up to now the largest protein assigned by solid-state NMR. Our results provide the basis to study high molecular weight proteins at atomic level. This is demonstrated in the second part with the first high-resolution solid-state NMR study of Ure2 and Sup35 prion fibrils. We describe the conformation of the functional domains and prion domains in the full-length fibrils and in isolation. The third fibrillar protein addressed in this work is the Parkinson’s disease related α-synuclein whereof we demonstrate the NMR resonance assignment and the secondary structure determination of a new polymorph. Thus, the studies described here provide new insights in the structural diversity of fibril architectures, and plead to view fibrils as individuals from a structural point of view, rather than a homogenous protein family
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Yao, Xuejun [Verfasser], Markus [Akademischer Betreuer] Zweckstetter, and Kai [Akademischer Betreuer] Tittmann. "Solution NMR-based characterization of the structure of the outer mitochondrial membrane protein Tom40 and a novel method for NMR resonance assignment of large intrinsically disordered proteins / Xuejun Yao. Gutachter: Markus Zweckstetter ; Kai Tittmann. Betreuer: Markus Zweckstetter." Göttingen : Niedersächsische Staats- und Universitätsbibliothek Göttingen, 2014. http://d-nb.info/1050873122/34.
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Sujeesh, K. S. "New NMR Methods with high resolution and sensitivity for rapid data analysis." Thesis, 2019. https://etd.iisc.ac.in/handle/2005/5041.
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work is focused on: (i) Development of new fast and sensitivity enhanced NMR methods in solution state and their application to metabolomics and (ii) development of novel NMR methods in solution state for the rapid resonance assignment of proteins. Described below are the projects carried out towards the dissertation of my PhD.
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Dubey, Abhinav. "Development Of NMR Methods For Metabolomics And Protein Resonance Assignments." Thesis, 2016. https://etd.iisc.ac.in/handle/2005/2633.
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Nuclear Magnetic Resonance (NMR) spectroscopy is a quantitative, non-invasive and non-destructive technique useful in biological studies. By manipulating the magnetization of nuclei with non-zero spin, NMR gives insights into atomic level details. Application of NMR as a tool for discovering structure, understanding dynamics of bio-molecules such as proteins, metabolites, DNA, RNA and their interactions constitutes the field of bio-molecular NMR. In this thesis, new methods for rapid data analysis of NMR spectrum of proteins and metabolites are proposed.
The first computational method, PROMEB (Pattern Recognition Based Assignment in Metabolomics) is useful for the identification and assignments of metabolites. This is an important step in metabolomics and is necessary for the discovery of new biomarkers. In NMR spectroscopy based studies, the conventional approach involves a database search, wherein chemical shifts are assigned to specific metabolites by use of a tolerance limit. This is inefficient because deviation in chemical shifts associated with pH or temperature variations, as well as missing peaks, impairs a robust comparison with the database. These drawbacks are overcome in PROMEB, which is a method based on matching the pattern of peaks of a metabolite in 2D [13C, 1H] HSQC NMR spectrum, rather than conventionally used absolute tolerance thresholds. A high success rate is obtained even in the presence of large chemical shift deviations such as 0.5 ppm in 1H and 3 ppm in 13C and missing peaks (up to 50%), compared to nearly no assignments obtained under these conditions with existing methods that employ a direct database search approach. The pattern recognition approach thus helps in identification and assignment of metabolites in-dependent of the pH, temperature, and ionic strength used, thereby obviating the need for spectral calibration with internal or external standards.
Another computational method, ChemSMP(Chemical Shifts to Metabolic Path-ways), is described which facilitates the identification of metabolic pathways from a single two dimensional (2D) NMR spectrum. Typically in other approaches, this is done after relevant metabolites are identified to allow their mapping onto specific metabolic pathways. This task is daunting due to the complex nature of cellular processes and the difficulty in establishing the identity of individual metabolites. ChemSMP uses a novel indexing and scoring system comprised of a uniqueness
score and a coverage score. Benchmarks show that ChemSMP has a positive prediction rate of > 90% in the presence of decluttered data and can sustain the same at 60 − 70% even in the presence of noise, such as deletions of peaks and chemical shift deviations. The method tested on NMR data acquired for a mixture of 20 amino acids shows a success rate of 93% in correct recovery of metabolic pathways.
The third method developed is a new approach for rapid resonance assignments in proteins based on amino acid selective unlabeling. The method involves choosing a set of multiple amino acid types for selective unlabeling and identifying specific tripeptides surrounding the labeled residues from specific 2D NMR spectra in a combinatorial manner. The methodology directly yields sequence specific resonance assignments, without requiring a contiguously assigned stretch of amino acid residues to be linked, and is applicable to deuterated proteins.
The fourth method involves a simple approach to rapidly identify amino acid types in proteins from a 2D NMR spectrum. The method is based on the fact that 13Cβ chemical shifts of different amino acid types fall in distinct spectral regions. By evolving the 13C chemical shifts in the conventional HNCACB or HN(CO)CACB type experiment for a single specified delay period, the phase of the cross peaks of different amino acid residues are modulated depending on their 13Cβ chemical shift values. Following this specified evolution period, the 2D HN projections of these experiments are acquired. The 13C evolution period can be chosen such that all residues belonging to a given set of amino acid types have the same phase pattern (positive or negative) facilitating their identification. This approach does not re-quire the preparation of any additional samples, involves the analysis of 2D [15N,1H] HSQC-type spectra obtained from the routinely used triple resonance experiments with minor modifications, and is applicable to deuterated proteins.
Finally, the practical application of these methods for laboratory research is presented. PROMEB and ChemSMP is used to study cancer cell metabolism in previously unexplored oncogenic cell line. PROMEB helped in assigning a differential metabolite present at high concentration in cancer cell line compared to control non-cancerous cell line. ChemSMP revealed active metabolic pathways responsible for regulating energy homeostasis of cancer cells which were previously reported in literature.
The two methods developed for rapid protein resonance assignments can be used in applications such as identifying active-site residues involved in ligand binding, phosphorylation, or protein-protein interactions. The phase modulated experiments will be useful for quick assignment of signals that shift during ligand binding or in combination with selective labeling/unlabeling approaches for identification of amino acid types to aid the sequential assignment process. Both the methodology was applied to two proteins: Ubiquitin (8 kDa) and L-IGFBP2 an intrinsically disordered protein (12 kDa), for demonstrating rapid resonance assignment using only set of 2D NMR experiments.
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Dubey, Abhinav. "Development Of NMR Methods For Metabolomics And Protein Resonance Assignments." Thesis, 2016. http://etd.iisc.ernet.in/handle/2005/2633.
Full textAbstract:
Nuclear Magnetic Resonance (NMR) spectroscopy is a quantitative, non-invasive and non-destructive technique useful in biological studies. By manipulating the magnetization of nuclei with non-zero spin, NMR gives insights into atomic level details. Application of NMR as a tool for discovering structure, understanding dynamics of bio-molecules such as proteins, metabolites, DNA, RNA and their interactions constitutes the field of bio-molecular NMR. In this thesis, new methods for rapid data analysis of NMR spectrum of proteins and metabolites are proposed.
The first computational method, PROMEB (Pattern Recognition Based Assignment in Metabolomics) is useful for the identification and assignments of metabolites. This is an important step in metabolomics and is necessary for the discovery of new biomarkers. In NMR spectroscopy based studies, the conventional approach involves a database search, wherein chemical shifts are assigned to specific metabolites by use of a tolerance limit. This is inefficient because deviation in chemical shifts associated with pH or temperature variations, as well as missing peaks, impairs a robust comparison with the database. These drawbacks are overcome in PROMEB, which is a method based on matching the pattern of peaks of a metabolite in 2D [13C, 1H] HSQC NMR spectrum, rather than conventionally used absolute tolerance thresholds. A high success rate is obtained even in the presence of large chemical shift deviations such as 0.5 ppm in 1H and 3 ppm in 13C and missing peaks (up to 50%), compared to nearly no assignments obtained under these conditions with existing methods that employ a direct database search approach. The pattern recognition approach thus helps in identification and assignment of metabolites in-dependent of the pH, temperature, and ionic strength used, thereby obviating the need for spectral calibration with internal or external standards.
Another computational method, ChemSMP(Chemical Shifts to Metabolic Path-ways), is described which facilitates the identification of metabolic pathways from a single two dimensional (2D) NMR spectrum. Typically in other approaches, this is done after relevant metabolites are identified to allow their mapping onto specific metabolic pathways. This task is daunting due to the complex nature of cellular processes and the difficulty in establishing the identity of individual metabolites. ChemSMP uses a novel indexing and scoring system comprised of a uniqueness
score and a coverage score. Benchmarks show that ChemSMP has a positive prediction rate of > 90% in the presence of decluttered data and can sustain the same at 60 − 70% even in the presence of noise, such as deletions of peaks and chemical shift deviations. The method tested on NMR data acquired for a mixture of 20 amino acids shows a success rate of 93% in correct recovery of metabolic pathways.
The third method developed is a new approach for rapid resonance assignments in proteins based on amino acid selective unlabeling. The method involves choosing a set of multiple amino acid types for selective unlabeling and identifying specific tripeptides surrounding the labeled residues from specific 2D NMR spectra in a combinatorial manner. The methodology directly yields sequence specific resonance assignments, without requiring a contiguously assigned stretch of amino acid residues to be linked, and is applicable to deuterated proteins.
The fourth method involves a simple approach to rapidly identify amino acid types in proteins from a 2D NMR spectrum. The method is based on the fact that 13Cβ chemical shifts of different amino acid types fall in distinct spectral regions. By evolving the 13C chemical shifts in the conventional HNCACB or HN(CO)CACB type experiment for a single specified delay period, the phase of the cross peaks of different amino acid residues are modulated depending on their 13Cβ chemical shift values. Following this specified evolution period, the 2D HN projections of these experiments are acquired. The 13C evolution period can be chosen such that all residues belonging to a given set of amino acid types have the same phase pattern (positive or negative) facilitating their identification. This approach does not re-quire the preparation of any additional samples, involves the analysis of 2D [15N,1H] HSQC-type spectra obtained from the routinely used triple resonance experiments with minor modifications, and is applicable to deuterated proteins.
Finally, the practical application of these methods for laboratory research is presented. PROMEB and ChemSMP is used to study cancer cell metabolism in previously unexplored oncogenic cell line. PROMEB helped in assigning a differential metabolite present at high concentration in cancer cell line compared to control non-cancerous cell line. ChemSMP revealed active metabolic pathways responsible for regulating energy homeostasis of cancer cells which were previously reported in literature.
The two methods developed for rapid protein resonance assignments can be used in applications such as identifying active-site residues involved in ligand binding, phosphorylation, or protein-protein interactions. The phase modulated experiments will be useful for quick assignment of signals that shift during ligand binding or in combination with selective labeling/unlabeling approaches for identification of amino acid types to aid the sequential assignment process. Both the methodology was applied to two proteins: Ubiquitin (8 kDa) and L-IGFBP2 an intrinsically disordered protein (12 kDa), for demonstrating rapid resonance assignment using only set of 2D NMR experiments.
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"Biomolecular solid-state nuclear magnetic resonance methods for spectral assignment and high-resolution structure determination of proteins." UNIVERSITY OF ILLINOIS AT URBANA-CHAMPAIGN, 2008. http://pqdtopen.proquest.com/#viewpdf?dispub=3301264.
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Franks, William Trent. "Biomolecular solid-state nuclear magnetic resonance methods for spectral assignment and high-resolution structure determination of proteins /." 2007. http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&res_dat=xri:pqdiss&rft_dat=xri:pqdiss:3301264.
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Thesis (Ph. D.)--University of Illinois at Urbana-Champaign, 2007.Source: Dissertation Abstracts International, Volume: 69-02, Section: B, page: 1027. Adviser: Chad M. Rienstra. Includes supplementary digital materials. Includes bibliographical references. Available on microfilm from Pro Quest Information and Learning.
Book chapters on the topic "Rapid resonance assignment of proteins"
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Robertson, Andrew D., and John L. Markley. "Methods of Proton Resonance Assignment for Proteins." In Biological Magnetic Resonance, 155–76. Boston, MA: Springer US, 1990. http://dx.doi.org/10.1007/978-1-4615-6549-9_4.
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Redfield, Christina, and James P. Robertson. "Assignment of the NMR Spectra of Homologous Proteins." In Computational Aspects of the Study of Biological Macromolecules by Nuclear Magnetic Resonance Spectroscopy, 303–16. Boston, MA: Springer US, 1991. http://dx.doi.org/10.1007/978-1-4757-9794-7_23.
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Kleywegt, Gerard J., Rolf Boelens, and Robert Kaptein. "STELLA and CLAIRE: A Seraglio of Programs for Human-Aided Assignment of 2D 1H NMR Spectra of Proteins." In Computational Aspects of the Study of Biological Macromolecules by Nuclear Magnetic Resonance Spectroscopy, 427–37. Boston, MA: Springer US, 1991. http://dx.doi.org/10.1007/978-1-4757-9794-7_34.
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Hiller, Sebastian, Christian Wasmer, Gerhard Wider, and Kurt Wüthrich. "Sequence-Specific Resonance Assignment of Soluble Nonglobular Proteins by 7D APSY-NMR Spectroscopy." In NMR with Biological Macromolecules in Solution, 72–77. WORLD SCIENTIFIC, 2021. http://dx.doi.org/10.1142/9789811235795_0008.
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Dijkgraaf, Ingrid, Stijn M. Agten, Matthias Bauwens, and Tilman M. Hackeng. "Strategies for Site-Specific Radiolabeling of Peptides and Proteins." In Radiopharmaceuticals [Working Title]. IntechOpen, 2021. http://dx.doi.org/10.5772/intechopen.99422.
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Although anatomical imaging modalities (X-ray, computed tomography (CT), magnetic resonance imaging (MRI)) still have a higher spatial resolution (0.1–1 mm) than molecular imaging modalities (single-photon emission computed tomography (SPECT), positron emission tomography (PET), optical imaging (OI)), the advantage of molecular imaging is that it can detect molecular and cellular changes at the onset of a disease before it leads to morphological tissue changes, which can be detected by anatomical imaging. During the last decades, noninvasive diagnostic imaging has encountered a rapid growth due to the development of dedicated imaging equipment for preclinical animal studies. In addition, the introduction of multimodality imaging (PET/CT, SPECT/CT, PET/MRI) which combines high-resolution conventional anatomical imaging with high sensitivity of tracer-based molecular imaging techniques has led to successful accomplishments in this exciting field. In this book chapter, we will focus on chemical synthesis techniques for site-specific incorporation of radionuclide chelators. Subsequently, radiolabeling based on complexation of a radionuclide with a chelator will be discussed, with focus on: diethylenetriaminepentaacetic acid (DTPA), 1,4,7,10-tetraazacyclododecane-tetraacetic acid (DOTA), 1,4,7-triazacyclononane-triacetic acid (NOTA), hexa-histidine (His-tag), and 6-hydrazinonicotinic acid (HYNIC) that allow the production of peptides labeled with 18F, 68Ga, 99mTc, and 111In – the currently most widely used isotopes.
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Kaplan, O., and J. S. Cohen. "Nuclear Magnetic Resonance Spectroscopy Studies of Cancer Cell Metabolism." In Biological NMR Spectroscopy. Oxford University Press, 1997. http://dx.doi.org/10.1093/oso/9780195094688.003.0030.
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Nuclear magnetic resonance spectroscopy (NMR) is a powerful technique that provides information on biochemical status and physiological processes both in-vitro and in-vivo. The metabolism of intact cells and tissues can be studied in a continuous manner, and thus, NMR is a unique non-invasive research tool enabling detection of the metabolic changes as they occur (Cohen et al., 1983; Morris, 1988; Daly and Cohen, 1989). The first NMR study of cellular metabolism was done some 20 years ago, when Moon and Richards reported on the diphosphoglyceric acid (DPG) and pH shifts in erythrocytes (Moon, and Richards, 1973). NMR studies of metabolism of tumor cells were initiated by Navon et al. who investigated phosphorylated compounds in Ehrlich ascites cells (Navon etal., 1977). The choice of the element and isotope for a specific study of metabolism depends on its NMR properties, and the required data. The proton has the highest NMR sensitivity, and is the most abundant nucleus in biological molecules. However, this may cause difficulties in the interpretation and assignment of the 1H NMR spectrum. Moreover, since metabolic studies are usually performed in aqueous solutions, the huge signal from the water protons should be suppressed. Similarly, the wide signals arising from proteins and membrane components should be suppressed. These problems can be addressed now by several innovative NMR methods (Daniels et al., 1976; van Zijl and Cohen, 1992). The most widely used nucleus in NMR studies of metabolism has been 31p (see reviews Cohen (1988); Kaplan et al. (1992)). Phosphorous NMR spectroscopy can provide data on energy metabolism and substrate utilization, phospholipid pathways, precise intracellular pH, and membrane permeability and ion and water distribution. The spectrum is easy to interpret, but the number of compounds which are detectable is limited. Carbon NMR is also useful for NMR studies of metabolism since it is found in most biological compounds; however, 13C has a natural abundance of only 1.1%, and 13C enrichment is necessary. Other nuclei which are used less often in NMR studies of cellular metabolism are 23Na (Gupta et al., 1984), 19F (Malet-Martino, et al., 1986), and rarely 15N (Legerton et al., 1983) and 39K (Brophy et al., 1983).
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McKeon, Andrew, and Nicholas L. Zalewski. "Rapid-Onset Hemibody Sensory Loss, Incoordination, and Muscle Jerking." In Mayo Clinic Cases in Neuroimmunology, edited by Andrew McKeon, B. Mark Keegan, and W. Oliver Tobin, 184–86. Oxford University Press, 2021. http://dx.doi.org/10.1093/med/9780197583425.003.0059.
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A 61-year-old woman with no pertinent medical history had progressive decline in multiple neurologic domains over the course of 2 months. She had development of progressive sensory loss in her left foot that subsequently spread up the left lower extremity and into the left upper extremity; accompanied by a sense of unsteadiness. Later, jerky movements of the left leg occurred while she was lying supine and sometimes when walking. At times, her left hand would wander involuntarily. Later in the course of her symptoms, mild short-term memory loss was also noted. On examination, she was unable to recall her home address, but findings were otherwise normal. She had mild gaze-evoked nystagmus and significant saccadic intrusion of smooth pursuits. A mild upper motor pattern of weakness, action myoclonus, hyperreflexia, and moderate loss of vibration was present on the left. Gait was markedly ataxic. Repeated magnetic resonance imaging of the brain 2 months after illness onset showed right parietal cortical hyperintense signal on diffusion-weighted imaging consistent with cortical ribboning, a common diagnostic finding early in the course of prion disease. Although characteristic of prion disease, similar imaging findings have been reported in autoimmune encephalitis and in the postictal setting. However, the putamen and caudate nucleus also demonstrated subtle asymmetric diffusion-weighted imaging hyperintense signal, which in the clinicoradiologic context is highly specific for prion disease. Electroencephalography showed frequent sharp wave discharges over right posterior temporal and left occipital head regions, along with frontal intermittent rhythmic delta slowing, consistent with encephalopathy (not otherwise specified). Real-time quaking-induced conversion testing of cerebrospinal fluid was positive for misfolded prion proteins. The positive real-time quaking-induced conversion result confirmed a diagnosis of sporadic Creutzfeldt-Jakob disease. The patient’s treatment was palliative. Hospice services implemented a home palliation program. Clonazepam was prescribed to reduce myoclonus. The patient died 18 weeks after onset of her neurologic symptoms. The differential diagnosis of a rapidly progressive multifocal neurologic syndrome includes many considerations but can be focused in complex situations by first confirming lesion localization and characterization with neuroimaging or other objective studies (eg, electromyography).
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Matthews, K. S., and R. Matthews. "Selective Chemical Deuteration of Aromatic Amino Acids: A Retrospective." In Biological NMR Spectroscopy. Oxford University Press, 1997. http://dx.doi.org/10.1093/oso/9780195094688.003.0021.
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In 1970 when we began post-doctoral work in the laboratory of Professor Oleg Jardetzky, selective deuteration of proteins to limit the number of protons present in the system for subsequent analysis was a newly developed and effective technique for NMR exploration of protein structure (Crespi et al., 1968; Markley et al., 1968). This approach allowed more facile assignment of specific resonances and generated the potential to follow the spectroscopic behavior of protons for a specific amino acid sidechain over a broad range of conditions. The primary method for labeling at that time involved growth of microorganisms (generally bacteria or algae) in D2O, followed by isolation of the deuteratedamino acids from a cellular protein hydrolysate. The amino acids isolated were, therefore, completely deuterated. Selective deuteration of a target protein was achieved by growing the producing organism on a mixture of completely deuterated and selected protonated amino acids under conditions that minimized metabolic interconversion of the amino acids. In one-dimensional spectra, aromatic amino acid resonances occur well downfield of the aliphatic resonances, and this region can therefore be examined somewhat independently by utilizing a single protonated aromatic amino acid to simplify the spectrum of the protein. However, the multiple spectral lines generated by aromatic amino acids can be complex and overlapping, precluding unequivocal interpretation. To address this complication, chemical methods were developed to both completely and selectively deuterate side chains of the aromatic amino acids, thereby avoiding the costly necessity of growing large volumes of microorganisms in D2O and subsequent tedious isolation procedures. In addition, selective deuteration of the amino acids simplified the resonance patterns and thereby facilitated assignment and interpretation of spectra. The methods employed were based on exchange phenomena reported in the literature and generated large quantities of material for use in growth of microorganisms for subsequent isolation of selectively labeled protein (Matthews et al., 1977a). The target protein for incorporation of the selectively deuterated aromatic amino acids generated by these chemical methods was the lactose repressor protein from Escherichia coli, and greatly simplified spectra of this 150,000 D protein were produced by this approach.
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Graves, Steven W., and John P. Nolan. "Molecular Assemblies, Probes, and Proteomics in Flow Cytometry." In Flow Cytometry for Biotechnology. Oxford University Press, 2005. http://dx.doi.org/10.1093/oso/9780195183146.003.0013.
Full textAbstract:
The many proteins and nucleic acids encoded in the genome predominantly perform their functions as macromolecular assemblies. In fact, modern biomedical research often targets the interactions of individual molecules of these assemblies, usually by disrupting or enhancing specific contacts, to provide treatment for many different diseases. Therefore, efficient pharmaceutical design requires knowledge of how macromolecular assemblies are built and function. To achieve this goal, sensitive and quantitative tools are essential. This chapter will discuss the use of flow cytometry as a general platform for sensitive measurement and quantification of molecular assemblies. First, this chapter will introduce general methods for analysis of molecular interactions along with a comparison of flow cytometry with these methods. Second, an overview of current flow cytometry instrumentation, assay technologies, and applications in molecular assembly analysis will be given. Third, the implementation of the above approaches in molecular assembly will be discussed. Finally, potential future directions of flow cytometry in molecular assembly analysis will be explored. At present, the analysis of macromolecular assemblies is performed by a wide variety of techniques that are chosen for the target molecules under study (proteins, DNA, lipids, etc.), the type of measurement required (kinetic or equilibrium), and whether the assembly of interest needs to be studied in vivo or in vitro. This continuum of techniques can be divided into the heterogeneous assays, which require a separation step to resolve products from reactants, and homogeneous assays, which can measure interactions without a separation step. Heterogeneous assays, in general, use radioisotopes, which are not perturbing; offer excellent sensitivity; and provide accurate quantification. The products are quantified after a separation step such as gel filtration, gel electrophoresis, or centrifugation. Rapid quench methods can provide subsecond kinetic resolution; however, the added separation steps are tedious and make collection of kinetic time courses difficult, as each time point must be separated and measured individually. Furthermore, in the time it takes the separation to occur, the interaction of interest can dissociate, which is a problem specific to low-affinity assemblies. Nonetheless, by using rapid chemical quench techniques, reaction times as short as a few milliseconds can be observed. Homogenous assays can be separated into solution- or surface-based assays. Solutionbased assays measure an optical signal generated by the assembly to quantify an interaction. High component concentrations (micromolar) allow changes in intrinsic molecular properties, such as protein fluorescence or circular dichroism, to be used to study molecular assemblies. For greater sensitivity (nanomole component concentrations), resonance energy transfer or polarization assays using exogenous fluorescent labels can be used. In combination with stopped-flow spectroscopy methodologies, solution-based assays allow reactions to monitored in a continuous fashion with submillisecond dead times.
Conference papers on the topic "Rapid resonance assignment of proteins"
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Champion, P. M., J. T. Sage, and P. Li. "Resonance Raman Studies of Electronic and Vibrational Relaxation Dynamics in Heme Proteins." In International Conference on Ultrafast Phenomena. Washington, D.C.: Optica Publishing Group, 1992. http://dx.doi.org/10.1364/up.1992.tuc6.
Full textAbstract:
We discuss the technique of resonance Raman saturation spectroscopy and present experimental results that probe relaxation processes in heme proteins following electronic excitation in the Soret band. The observable relaxation time scales are limited by the laser excitation rate, kL, rather than by the laser pulse width (~10 ns). Analysis of the data using a model that allows for parallel relaxation channels leads to effective electronic relaxation times for deoxymyoglobin (2ps), ferrocytochrome c (3ps), and deoxyhemoglobin (1ps). The more rapid relaxation times found for hemoglobin and its isolated chains are not currently understood. The Raman depolarization ratio is predicted to increase at high laser flux, due to the preparation of a partially oriented sample by photoselective excitation. Such effects are observed in heme systems and the relaxation times extracted from the depolarization analysis are in agreement with the measurements of Raman intensity saturation. Studies of the asymmetric broadening of the ν 4 mode of cytochrome c at high laser flux, reveal that the lineshapes in the Stokes and anti-Stokes region are inequivalent. Time-reversal symmetry dictates that this broadening is due to an underlying Raman band, associated with an excited electronic state that is populated at high laser flux. Similar linebroadening effects, observed in hemoglobin and myoglobin samples, are also shown to arise from Raman scattering of excited electronic states rather than Rabi broadening1 or anharmonic coupling to vibrationally hot low frequency modes2.
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Furis, B. C., M. J. Jorgensem, M. J. Rabiet, A. B. Contor, C. L. Brown, C. B. Shoemaker, and B. Furie. "RECOGNITION SITE DIRECTING GAMMA-CARBOXYLATION RESIDES ON THE PROPEPTIDES OF FACTOR IX AND PROTRROMBIN." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1643992.
Full textAbstract:
Factor IX and prothrombin vitamin K-dependent proteins that participate in blood coagulation undergo post-translationalmodification in which glutamic acid residues in the amino terminus of the protein are converted to gamma-carboxyglutamic acid residues. This modification confers divalent metal ion binding ability upon the proteins.As a consequence of binding divalent metal ions these proteins undergoconformational changes necessary for biological function.The vitamin K-dependent proteins are synthesized with an NH2-terminal extension. The region distal to the NH2-terminus of the mature protein is a prototypic signal sequence while the proximal region is a propeptide with homology among the vitamin K-dependent proteins. The boundary between the pre and pro sequences has been established for factor IX by analysis of three naturally occurring factor IX mutants factor IX Cambridge factor IX Oxford-3 and factor IX San Dimas, in which processing is incomplete.For human factor IX the propeptide extends from residue -18 to -1. The homology among the propeptides of vitamin K-dependent proteins suggests that the propeptide may designate adjacent gamma-carboxyglutamic acids for carboxylation. To test this hypothesis alterations in sequence were introduced into the propeptide region of human factor IX cDNA by oligonucleotide directed site specific mutagenesis.Mutated genes were expressed in Chinese hamster ovary cells. Rapid and efficient isolationof the mutant proteins by immunoaffinity chromatography permitted detailed analysis of the mutants on quantities of protein easily obtainable at low expression levels. The extent of gamma-carboxylation was assessed by the ability of the mutant proteins to interact with conformation specific antibodies directed against the gamma-carboxyglutamic acid-dependent metal stabilized native structure of factor IX as well as by direct amino acid analysis. Unmodified recombinant factor IX contained, on average, 9 gamma-carboxyglutamic acid residues, as compared to 12 for plasma factor IX. About 70% of the recombinant wild type factor IX bound to the conformation specific antibodies. Deletion of the propiece or point mutations at residues -10 or -16 led to secretion of uncarboxylated factor IX unreaotive with antibodies specific for the native structure but with the NH2-terminus of mature factor IX. In order to assess the universality of these observations we have recently cloned human prothrombin cDNA and expressed the gene in the same Chinese hamster ovary cell system used for factor IX. In contrast to factor IX, at low levels ofexpressionof the prothrombin gene, the prothrombin is fully carboxylated relative to a plasma prothrombin standard.The recombinant prothrombin exhibits the same specific clotting activity as plasma derivedprothrombin and is fully native as evaluated by conformation specific antibodies. At high levels of expression the capacityof the cells to carboxylate prothrombin can be exceeded leading to secretion of under carboxylated prothrombin. However, the absolute amount of fully carboxylated prothrombin that can be produced in this system appears to be a least fivefold greater that the absolute amount of highly carboxylated factor IX that can be synthesized.The elimination of carboxylation observed upon mutation of the propiece of factor IX suggest that the propiece contains a recognition element required for carboxylation of the protein. Assignment of a functional role to the propiece of factor IX represents the first determination of function for any pro sequence. It is anticipated that extension of these studies to prothrombin will demonstrate that this recognition signal is used by all the members of this class of proteins. In order to determine if the propiece is sufficient to designate a protein for gamma-carboxylation we are currently constructing chimeric proteins incorporating the propieceof prothrombin into the cDNA of normally uncarboxylated proteins.
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McCarthy, Michael C., and R. W. Field. "Application of a sideband oodr Zeeman spectroscopy to diatomic molecules." In OSA Annual Meeting. Washington, D.C.: Optica Publishing Group, 1990. http://dx.doi.org/10.1364/oam.1990.wn2.
Full textAbstract:
In order to test simple atom-in-molecule concepts in the theory of the electronic structure of transition metal monohydrides and monohalides it is necessary to extract key electronic properties from the spectra of these molecules. Unfortunately, the spectra of diatomic molecules containing an atom with a partially filled d-shell are complex. Consequently, it is desirable to develop spectroscopic techniques and strategies whereby spectra can be simplified and rapid generation of diagnostically important spectral information is possible. Sideband optical-optical double resonance-Zeeman spectroscopy (SOODRZ) is one such technique. SOODRZ is one-laser, two-color, absorption-based technique which presents sub-Doppler Zeeman splittings in a simple, diagnostically useful format. Unlike other sub-Doppler techniques where complex Zeeman splitting patterns obscure the tuning coefficients of either the upper state or lower state, in SOODRZ, only two Zeeman resonance features (single upper and single lower state resonance) are observed. The Zeeman information (J, Q-dependent Zeeman tuning coefficients) can then be used for rotational and electronic assignment, we will illustrate the usefulness of SOODRZ by showing examples from the electronic spectrum of NiH. We will also discuss the lineshape and sensitivity of SOODRZ for a variety of detection schemes using both electro-optic and acousto-optic modulators.