The reaction cycle and the major structural states of the molecular chaperone GroEL and its cochaperone, GroES, are well characterized. In contrast, very little is known about the nonnative states of the substrate polypeptide acted on by the chaperonin machinery. In this study, we investigated the substrate protein human dihydrofolate reductase (hDHFR) while bound to GroEL or to a single-ring analog, SR1, by NMR spectroscopy in solution under conditions where hDHFR was efficiently recovered as a folded, enzymatically active protein from the stable complexes upon addition of ATP and GroES. By using the NMR techniques of transverse relaxation-optimized spectroscopy (TROSY), cross-correlated relaxation-induced polarization transfer (CRIPT), and cross-correlated relaxation-enhanced polarization transfer (CRINEPT), bound hDHFR could be observed directly. Measurements of the buildup of hDHFR NMR signals by different magnetization transfer mechanisms were used to characterize the dynamic properties of the NMR-observable parts of the bound substrate. The NMR data suggest that the bound state includes random coil conformations devoid of stable native-like tertiary contacts and that the bound hDHFR might best be described as a dynamic ensemble of randomly structured conformers.
Influence of substrate modification and C-terminal truncation on the active site structure of substrate-bound heme oxygenase from Neisseriae meningitidis; A 1H NMR study.
Influence of substrate modification and C-terminal truncation on the active site structure of substrate-bound heme oxygenase from Neisseriae meningitidis; A 1H NMR study.
Influence of substrate modification and C-terminal truncation on the active site structure of substrate-bound heme oxygenase from Neisseriae meningitidis; A 1H NMR study.
Biochemistry. 2011 Aug 27;
Authors: Peng D, Satterlee JD, Ma LH, Dallas JL, Smith KM, Zhang X, Sato M, La Mar GN
Abstract
Heme oxygenase, HO, from the pathogenic bacterium N. meningitidis, NmHO, which...
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NMR spectroscopy with the stringent substrate rhodanese bound to the single-ring variant SR1 of the E. coli chaperonin GroEL.
NMR spectroscopy with the stringent substrate rhodanese bound to the single-ring variant SR1 of the E. coli chaperonin GroEL.
NMR spectroscopy with the stringent substrate rhodanese bound to the single-ring variant SR1 of the E. coli chaperonin GroEL.
Protein Sci. 2011 Jun 1;
Authors: Koculi E, Horst R, Horwich AL, Wüthrich K
NMR observation of the uniformly (2) H,(15) N-labeled stringent 33 kDa substrate protein rhodanese in a productive complex with the uniformly (14) N-labeled 400 kDa single-ring version of the E. coli chaperonin GroEL, SR1,...
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[NMR paper] Direct observation and characterization of DMPC/DHPC aggregates under conditions rele
Direct observation and characterization of DMPC/DHPC aggregates under conditions relevant for biological solution NMR.
Related Articles Direct observation and characterization of DMPC/DHPC aggregates under conditions relevant for biological solution NMR.
Biochim Biophys Acta. 2004 Aug 30;1664(2):241-56
Authors: van Dam L, Karlsson G, Edwards K
We have used cryo-transmission electron microscopy (cryo-TEM) for inspection of aggregates formed by dimyristoylphosphatidylcholine (DMPC) and dihexanoylphosphatidylcholine (DHPC) in aqueous solution at...
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[NMR paper] Multiple cycles of global unfolding of GroEL-bound cyclophilin A evidenced by NMR.
Multiple cycles of global unfolding of GroEL-bound cyclophilin A evidenced by NMR.
http://www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--linkinghub.elsevier.com-ihub-images-PubMedLink.gif Related Articles Multiple cycles of global unfolding of GroEL-bound cyclophilin A evidenced by NMR.
J Mol Biol. 1997 Sep 5;271(5):803-18
Authors: Nieba-Axmann SE, Ottiger M, Wüthrich K, Plückthun A
GroE, the chaperonin system of Escherichia coli, prevents the aggregation of partially folded or misfolded proteins by complexing them in a form competent for...
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08-22-2010 05:08 PM
[NMR paper] NMR observation of substrate in the binding site of an active sugar-H+ symport protei
NMR observation of substrate in the binding site of an active sugar-H+ symport protein in native membranes.
http://www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--www.pubmedcentral.nih.gov-corehtml-pmc-pmcgifs-pubmed-pmc.gif Related Articles NMR observation of substrate in the binding site of an active sugar-H+ symport protein in native membranes.
Proc Natl Acad Sci U S A. 1994 Apr 26;91(9):3877-81
Authors: Spooner PJ, Rutherford NG, Watts A, Henderson PJ
NMR methods have been adopted to observe directly the characteristics of substrate...
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[NMR paper] NMR observation of substrate in the binding site of an active sugar-H+ symport protei
NMR observation of substrate in the binding site of an active sugar-H+ symport protein in native membranes.
http://www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--www.pubmedcentral.nih.gov-corehtml-pmc-pmcgifs-pubmed-pmc.gif Related Articles NMR observation of substrate in the binding site of an active sugar-H+ symport protein in native membranes.
Proc Natl Acad Sci U S A. 1994 Apr 26;91(9):3877-81
Authors: Spooner PJ, Rutherford NG, Watts A, Henderson PJ
NMR methods have been adopted to observe directly the characteristics of substrate...
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08-22-2010 03:33 AM
[NMR paper] Direct observation and elucidation of the structures of aged and nonaged phosphorylat
Direct observation and elucidation of the structures of aged and nonaged phosphorylated cholinesterases by 31P NMR spectroscopy.
Related Articles Direct observation and elucidation of the structures of aged and nonaged phosphorylated cholinesterases by 31P NMR spectroscopy.
Biochemistry. 1993 Dec 14;32(49):13441-50
Authors: Segall Y, Waysbort D, Barak D, Ariel N, Doctor BP, Grunwald J, Ashani Y
31P NMR spectroscopy of butyrylcholinesterase (BChE), acetylcholinesterase (AChE), and chymotrypsin (Cht) inhibited by pinacolyl...