Related ArticlesAn advantage for use of isotope labeling and NMR chemical shifts to analyze the structure of four homologous IgG-binding domains of staphylococcal protein A.
J Biochem Biophys Methods. 2000 Jan 3;42(1-2):35-47
Because of the complexity arising from the large molecular size and the amino acid sequence homologies of IgG-binding domains of Staphylococcal Protein A (SpA), we have introduced, a combination of stable isotope labeling and both qualitative and quantitative investigations of the structural dependence of the NMR chemical shifts for its structure analysis. In order to enable selective isotope labeling with high efficiency, a mutated low molecular weight Protein A (LPA; MWt = 27 kDa) which consists of E, D, A, B and 13 residues of the C-domain was used in this study. Amide proton chemical shifts, measured using uniformly 15N-labeled LPA and LPA labeled selectively with 15N-alanine, show that the turn between helices 1 and 2, and its tertiary interactions with helix 3, are very similar in all domains. This contradicts previous results obtained using independent structure calculations on isolated domains. The close similarity in NH and 15N chemical shifts of alanine residues in the interdomain linker suggests that the linker maintains a similar structure both in isolated domains and in the intact protein. We show that the high-field shifted methyl signal of Ala 48 is affected by the ring-current effect arising from Phe 30, and has a very similar helical environment in all four domains. Thus, helix 3 is present in all domains, as we previously reported [Kikuchi et al., J Biochem Biophys Method, 1999:38:203-208], even though it is not observed in the crystal structure [Deisenhofer J. Biochemistry 1981;20:2361-2370].
Isotope labeling strategies for NMR studies of RNA
Isotope labeling strategies for NMR studies of RNA
Abstract The known biological functions of RNA have expanded in recent years and now include gene regulation, maintenance of sub-cellular structure, and catalysis, in addition to propagation of genetic information. As for proteins, RNA function is tightly correlated with structure. Unlike proteins, structural information for larger, biologically functional RNAs is relatively limited. NMR signal degeneracy, relaxation problems, and a paucity of long-range 1Hâ??1H dipolar contacts have limited the utility of traditional NMR approaches....
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Application of SAIL phenylalanine and tyrosine with alternative isotope-labeling patterns for protein structure determination
Application of SAIL phenylalanine and tyrosine with alternative isotope-labeling patterns for protein structure determination
Abstract The extensive collection of NOE constraint data involving the aromatic ring signals is essential for accurate protein structure determination, although it is often hampered in practice by the pervasive signal overlapping and tight spin couplings for aromatic rings. We have prepared various types of stereo-array isotope labeled phenylalanines (ε- and Îĥ-SAIL Phe) and tyrosine (ε-SAIL Tyr) to overcome these problems (Torizawa et al. 2005), and proven...
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[NMR paper] Combining inducible protein overexpression with NMR-grade triple isotope labeling in the cyanobacterium Anabaena sp. PCC 7120.
Combining inducible protein overexpression with NMR-grade triple isotope labeling in the cyanobacterium Anabaena sp. PCC 7120.
Related Articles Combining inducible protein overexpression with NMR-grade triple isotope labeling in the cyanobacterium Anabaena sp. PCC 7120.
Biotechniques. 2005 Sep;39(3):405-11
Authors: Desplancq D, Bernard C, Sibler AP, Kieffer B, Miguet L, Potier N, Van Dorsselaer A, Weiss E
The difficulty and expense of preparing protein samples highly enriched in stable isotopes is a bottleneck for structural studies by nuclear...
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12-01-2010 06:56 PM
[NMR paper] Expression, purification, and isotope labeling of a gp120 V3 peptide and production o
Expression, purification, and isotope labeling of a gp120 V3 peptide and production of a Fab from a HIV-1 neutralizing antibody for NMR studies.
Related Articles Expression, purification, and isotope labeling of a gp120 V3 peptide and production of a Fab from a HIV-1 neutralizing antibody for NMR studies.
Protein Expr Purif. 2002 Apr;24(3):374-83
Authors: Sharon M, Görlach M, Levy R, Hayek Y, Anglister J
Most human immunodeficiency virus type 1 (HIV-1) neutralizing antibodies in infected individuals and in immunized animals are directed...
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11-24-2010 08:49 PM
[NMR paper] New developments in isotope labeling strategies for protein solution NMR spectroscopy
New developments in isotope labeling strategies for protein solution NMR spectroscopy.
Related Articles New developments in isotope labeling strategies for protein solution NMR spectroscopy.
Curr Opin Struct Biol. 2000 Oct;10(5):585-92
Authors: Goto NK, Kay LE
The development of novel isotope labeling strategies for proteins has facilitated the study of the structure and dynamics of these molecules. In addition, the recent emergence of alternative methods of bacterial expression for obtaining isotopically labeled proteins permits the study of...
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11-19-2010 08:29 PM
[NMR paper] Cell-free synthesis and amino acid-selective stable isotope labeling of proteins for
Cell-free synthesis and amino acid-selective stable isotope labeling of proteins for NMR analysis.
Related Articles Cell-free synthesis and amino acid-selective stable isotope labeling of proteins for NMR analysis.
J Biomol NMR. 1995 Sep;6(2):129-34
Authors: Kigawa T, Muto Y, Yokoyama S
For the application of multidimensional NMR spectroscopy to larger proteins, it would be useful to perform selective labeling of one of the 20 amino acids. For some amino acids, however, amino acid metabolism drastically reduces the efficiency and selectivity...
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08-22-2010 03:50 AM
[NMR paper] Uniform 13C isotope labeling of proteins with sodium acetate for NMR studies: applica
Uniform 13C isotope labeling of proteins with sodium acetate for NMR studies: application to human carbonic anhydrase II.
Related Articles Uniform 13C isotope labeling of proteins with sodium acetate for NMR studies: application to human carbonic anhydrase II.
Biochemistry. 1991 May 7;30(18):4491-4
Authors: Venters RA, Calderone TL, Spicer LD, Fierke CA
Uniform double labeling of proteins for NMR studies can be prohibitively expensive, even with an efficient expression and purification scheme, due largely to the high cost of glucose. We...
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08-21-2010 11:16 PM
A simple method for amino acid selective isotope labeling of recombinant proteins in E. coli
A simple method for amino acid selective isotope labeling of recombinant proteins in E. coli
Kit I. Tong, Masayuki Yamamoto and Toshiyuki Tanaka
Journal of Biomolecular NMR; 2008; 42(1); pp 59-67
Abstract:
A simple and user-friendly method of labeling protein selectively with amino acids in vivo is introduced. This technique does not require the use of transaminase-deficient or auxotrophic strains. By manipulating the product feedback inhibitory loops of the E. coli amino acid metabolic pathways and, if necessary, by using enzyme inhibitors, proteins were labeled efficiently in vivo...
An advantage for use of isotope labeling and NMR chemical shifts to analyze the struc
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