Related ArticlesDeterminants of protein hyperthermostability: purification and amino acid sequence of rubredoxin from the hyperthermophilic archaebacterium Pyrococcus furiosus and secondary structure of the zinc adduct by NMR.
Biochemistry. 1991 Nov 12;30(45):10885-95
Authors: Blake PR, Park JB, Bryant FO, Aono S, Magnuson JK, Eccleston E, Howard JB, Summers MF, Adams MW
The purification, amino acid sequence, and two-dimensional 1H NMR results are reported for the rubredoxin (Rd) from the hyperthermophilic archaebacterium Pyrococcus furiosus, an organism that grows optimally at 100 degrees C. The molecular mass (5397 Da), iron content (1.2 +/- 0.2 g-atom of Fe/mol), UV-vis spectrophotometric properties, and amino acid sequence (60% sequence identity with Clostridium pasteurianum Rd) are found to be typical of this class of redox protein. However, P. furiosus Rd is remarkably thermostable, being unaffected after incubation for 24 h at 95 degrees C. One- and two-dimensional 1H nuclear magnetic resonance spectra of the oxidized [Fe(III)Rd] and reduced [Fe(II)Rd] forms of P. furiosus Rd exhibited substantial paramagnetic line broadening, and this precluded detailed 3D structural studies. The apoprotein was not readily amenable to NMR studies due to apparent protein oxidation involving the free cysteine sulfhydryls. However, high-quality NMR spectra were obtained for the Zn-substituted protein, Zn(Rd), enabling detailed NMR signal assignment for all backbone amide and alpha and most side-chain protons. Secondary structural elements were determined from qualitative analysis of 2D Overhauser effect spectra. Residues A1-K6, Y10-E14, and F48-E51 form a three-strand antiparallel beta-sheet, which comprises ca. 30% of the primary sequence. Residues C5-Y10 and C38-A43 form types I and II amide-sulfur tight turns common to iron-sulfur proteins. These structural elements are similar to those observed by X-ray crystallography for native Rd from the mesophile C. pasteurianum. However, the beta-sheet domain in P. furiosus Rd is larger than that in C. pasteurianum Rd and appears to begin at the N-terminal residue. From analysis of the secondary structure, potentially stabilizing electrostatic interactions involving the charged groups of residues Ala(1), Glu(14), and Glu(52) are proposed. These interactions, which are not present in rubredoxins from mesophilic organisms, may prevent the beta-sheet from "unzipping" at elevated temperatures.
Amino acid selective unlabeling for sequence specific resonance assignments in proteins
Amino acid selective unlabeling for sequence specific resonance assignments in proteins
Abstract Sequence specific resonance assignment constitutes an important step towards high-resolution structure determination of proteins by NMR and is aided by selective identification and assignment of amino acid types. The traditional approach to selective labeling yields only the chemical shifts of the particular amino acid being selected and does not help in establishing a link between adjacent residues along the polypeptide chain, which is important for sequential assignments. An alternative...
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Generation of Pseudocontact Shifts in Protein NMR Spectra with a Genetically Encoded Cobalt(II)-Binding Amino Acid.
Generation of Pseudocontact Shifts in Protein NMR Spectra with a Genetically Encoded Cobalt(II)-Binding Amino Acid.
Generation of Pseudocontact Shifts in Protein NMR Spectra with a Genetically Encoded Cobalt(II)-Binding Amino Acid.
Angew Chem Int Ed Engl. 2011 Jan 17;50(3):692-4
Authors: Nguyen TH, Ozawa K, Stanton-Cook M, Barrow R, Huber T, Otting G
[NMR paper] Site-selective screening by NMR spectroscopy with labeled amino acid pairs.
Site-selective screening by NMR spectroscopy with labeled amino acid pairs.
Related Articles Site-selective screening by NMR spectroscopy with labeled amino acid pairs.
J Am Chem Soc. 2002 Mar 20;124(11):2446-7
Authors: Weigelt J, van Dongen M, Uppenberg J, Schultz J, Wikström M
A new method for site-selective screening by NMR is presented. The core of the new method is the dual amino acid sequence specific labeling technique. Amino acid X is labeled with (13)C and amino acid Y is labeled with (15)N. Provided only one XY pair occurs in the...
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[NMR paper] Amino acid-specific isotopic labeling and active site NMR studies of iron(II)- and ir
Amino acid-specific isotopic labeling and active site NMR studies of iron(II)- and iron(III)-superoxide dismutase from Escherichia coli.
Related Articles Amino acid-specific isotopic labeling and active site NMR studies of iron(II)- and iron(III)-superoxide dismutase from Escherichia coli.
J Biomol NMR. 2000 Aug;17(4):311-22
Authors: Sorkin DL, Miller AF
We have developed and employed multiple amino acid-specific isotopic labeling schemes to obtain definitive assignments for active site 1H NMR resonances of iron(II)- and iron(III)-superoxide...
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[NMR paper] Glutaredoxin-3 from Escherichia coli. Amino acid sequence, 1H AND 15N NMR assignments
Glutaredoxin-3 from Escherichia coli. Amino acid sequence, 1H AND 15N NMR assignments, and structural analysis.
http://www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--highwire.stanford.edu-icons-externalservices-pubmed-standard-jbc_full_free.gif Related Articles Glutaredoxin-3 from Escherichia coli. Amino acid sequence, 1H AND 15N NMR assignments, and structural analysis.
J Biol Chem. 1996 Mar 22;271(12):6736-45
Authors: Aslund F, Nordstrand K, Berndt KD, Nikkola M, Bergman T, Ponstingl H, Jörnvall H, Otting G, Holmgren A
The primary and...
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08-22-2010 02:27 PM
[NMR paper] Determinants of protein hyperthermostability: purification and amino acid sequence of
Determinants of protein hyperthermostability: purification and amino acid sequence of rubredoxin from the hyperthermophilic archaebacterium Pyrococcus furiosus and secondary structure of the zinc adduct by NMR.
Related Articles Determinants of protein hyperthermostability: purification and amino acid sequence of rubredoxin from the hyperthermophilic archaebacterium Pyrococcus furiosus and secondary structure of the zinc adduct by NMR.
Biochemistry. 1991 Nov 12;30(45):10885-95
Authors: Blake PR, Park JB, Bryant FO, Aono S, Magnuson JK, Eccleston E, Howard...
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[NMR paper] Application of amino acid type-specific 1H- and 14N-labeling in a 2H-, 15N-labeled ba
Application of amino acid type-specific 1H- and 14N-labeling in a 2H-, 15N-labeled background to a 47 kDa homodimer: potential for NMR structure determination of large proteins.
Related Articles Application of amino acid type-specific 1H- and 14N-labeling in a 2H-, 15N-labeled background to a 47 kDa homodimer: potential for NMR structure determination of large proteins.
J Biomol NMR. 1999 May;14(1):79-83
Authors: Kelly MJ, Krieger C, Ball LJ, Yu Y, Richter G, Schmieder P, Bacher A, Oschkinat H
NMR investigations of larger macromolecules (> 20...