[NMR paper] Disassembly/reassembly strategy for the production of highly pure GroEL, a tetradecameric supramolecular machine, suitable for quantitative NMR, EPR and mutational studies.
Disassembly/reassembly strategy for the production of highly pure GroEL, a tetradecameric supramolecular machine, suitable for quantitative NMR, EPR and mutational studies.
Related ArticlesDisassembly/reassembly strategy for the production of highly pure GroEL, a tetradecameric supramolecular machine, suitable for quantitative NMR, EPR and mutational studies.
Protein Expr Purif. 2017 Sep 22;:
Authors: Wälti MA, Clore GM
Abstract
GroEL, a prototypical member of the chaperonin class of chaperones, is a large supramocular machine that assists protein folding and plays an important role in proteostasis. GroEL comprises two heptameric rings, each of which encloses a large cavity that provides a folding chamber for protein substrates. Many questions remain regarding the mechanistic details of GroEL facilitated protein folding. Thus, data at atomic resolution of the type provided by NMR and EPR are invaluable. Such studies often require complete deuteration of GroEL, uniform or residue specific (13)C and (15)N isotope labeling, and the introduction of selective cysteine mutations for site-specific spin labeling. In addition, high purity GroEL is essential for detailed studies of substrate-GroEL interactions as quantitative interpretation is impossible if the cavities are already occupied and blocked by other protein substrates present in the bacterial expression system. Here we present a new purification protocol designed to provide highly pure GroEL devoid of non-specific protein substrate contamination.
PMID: 28951283 [PubMed - as supplied by publisher]
Synthesis,Isotopic Enrichment, and Solid-State NMRCharacterization of Zeolites Derived from the Assembly, Disassembly,Organization, Reassembly Process
Synthesis,Isotopic Enrichment, and Solid-State NMRCharacterization of Zeolites Derived from the Assembly, Disassembly,Organization, Reassembly Process
Giulia P. M. Bignami, Daniel M. Dawson, Valerie R. Seymour, Paul S. Wheatley, Russell E. Morris and Sharon E. Ashbrook
http://pubs.acs.org/appl/literatum/publisher/achs/journals/content/jacsat/0/jacsat.ahead-of-print/jacs.7b00386/20170329/images/medium/ja-2017-00386n_0012.gif
Journal of the American Chemical Society
DOI: 10.1021/jacs.7b00386
http://feeds.feedburner.com/~ff/acs/jacsat?d=yIl2AUoC8zA...
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A systematic mutagenesis-driven strategy for site-resolved NMR studies of supramolecular assemblies
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Abstract Obtaining sequence-specific assignments remains a major bottleneck in solution NMR investigations of supramolecular structure, dynamics and interactions. Here we demonstrate that resonance assignment of methyl probes in high molecular weight protein assemblies can be efficiently achieved by combining fast NMR experiments, residue-type-specific isotope-labeling and automated site-directed mutagenesis. The utility of this general and straightforward strategy is demonstrated through...
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A systematic mutagenesis-driven strategy for site-resolved NMR studies of supramolecular assemblies.
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Authors: Amero C, Asunción Durá M, Noirclerc-Savoye M, Perollier A, Gallet B, Plevin MJ, Vernet T, Franzetti B, Boisbouvier J
Obtaining sequence-specific assignments remains a major bottleneck in solution NMR investigations of supramolecular structure, dynamics and interactions. Here we demonstrate that resonance...
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10-20-2010 06:50 AM
Strategy for complete NMR assignment of disordered proteins with highly repetitive se
Strategy for complete NMR assignment of disordered proteins with highly repetitive sequences based on resolution-enhanced 5D experiments
Abstract A strategy for complete backbone and side-chain resonance assignment of disordered proteins with highly repetitive sequence is presented. The protocol is based on three resolution-enhanced NMR experiments: 5D HN(CA)CONH provides sequential connectivity, 5D HabCabCONH is utilized to identify amino acid types, and 5D HC(CC-TOCSY)CONH is used to assign the side-chain resonances. The improved resolution was achieved by a combination of high...
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10-06-2010 02:16 AM
Strategy for complete NMR assignment of disordered proteins with highly repetitive se
Strategy for complete NMR assignment of disordered proteins with highly repetitive sequences based on resolution-enhanced 5D experiments.
Strategy for complete NMR assignment of disordered proteins with highly repetitive sequences based on resolution-enhanced 5D experiments.
J Biomol NMR. 2010 Oct 2;
Authors: MotáÄ?ková V, NováÄ?ek J, Zawadzka-Kazimierczuk A, Kazimierczuk K, ZÃ*dek L, Sanderová H, Krásný L, KoźmiÅ?ski W, SklenáÅ? V
A strategy for complete backbone and side-chain resonance assignment of disordered proteins with highly...
Disassembly/reassembly strategy for the production of highly pure GroEL, a tetradecameric supramolecular machine, suitable for quantitative NMR, EPR and mutational studies.
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