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Title: Four-dimensional sup 13 C/ sup 13 C-edited nuclear Overhauser Enhancement Spectroscopy of a protein in solution: Application to interleukin 1. beta

Abstract

A four-dimensional {sup 13}C/{sup 13}C-edited NOESY experiment is described which dramatically improves the resolution of protein NMR spectra and enables the straightforward assignment of nuclear Overhauser effects involving aliphatic and/or aromatic protons in larger proteins. The experiment is demonstrated for uniformly (>95{percent}) {sup 13}C-labeled interleukin 1{beta}, a protein of 153 residues and 17.4 kDa, which plays a key role in the immune response. NOEs between aliphatic and/or aromatic protons are first spread out into a third dimension by the {sup 13}C chemical shift of the carbon atom attached to the originating proton and subsequently into a fourth dimension by the {sup 13}C chemical shift of the carbon atom attached to the destination proton. Thus, each NOE cross peak is labeled by four chemical shifts. By this means, ambiguities in the assignment of NOEs that arise from chemical shift overlap and degeneracy are completely removed. Further, NOEs between protons with the same chemical shifts can readily be detected providing their attached carbon atoms have different {sup 13}C chemical shifts. The design of the pulse sequence requires special care to minimize the level of artifacts arising from undesired coherence transfer pathways, and in particular those associated with diagonal peaks which correspond tomore » magnetization that has not been transferred from one proton to another. The 4D {sup 13}C/{sup 13}C-edited NOESY experiment is characterized by high sensitivity as the through-bond transfer steps involve the large {sup 1}J{sub CH} (130 Hz) couplings, and it is possible to obtain high-quality spectra on 1-2 mM samples of {sup 13}C-labeled protein in as little as 3 days. This experiment should open up the application of protein structure determination by NMR to a large number of medium-sized proteins (150-300 residues) of biological interest.« less

Authors:
; ; ;  [1]
  1. (National Institutes of Health, Bethesda, MD (USA))
Publication Date:
OSTI Identifier:
5613529
Resource Type:
Journal Article
Journal Name:
Biochemistry; (USA)
Additional Journal Information:
Journal Volume: 30:1; Journal ID: ISSN 0006-2960
Country of Publication:
United States
Language:
English
Subject:
62 RADIOLOGY AND NUCLEAR MEDICINE; LYMPHOKINES; NUCLEAR MAGNETIC RESONANCE; PROTEIN STRUCTURE; GEOMETRY; AQUEOUS SOLUTIONS; CARBON 13; CHEMICAL SHIFT; ESCHERICHIA COLI; IMMUNE REACTIONS; OVERHAUSER EFFECT; BACTERIA; CARBON ISOTOPES; DISPERSIONS; EVEN-ODD NUCLEI; GROWTH FACTORS; ISOTOPES; LIGHT NUCLEI; MAGNETIC RESONANCE; MATHEMATICS; MICROORGANISMS; MITOGENS; MIXTURES; NUCLEI; ORGANIC COMPOUNDS; PROTEINS; RESONANCE; SOLUTIONS; STABLE ISOTOPES; 550601* - Medicine- Unsealed Radionuclides in Diagnostics

Citation Formats

Clore, G.M., Kay, L.E., Bax, A., and Gronenborn, A.M. Four-dimensional sup 13 C/ sup 13 C-edited nuclear Overhauser Enhancement Spectroscopy of a protein in solution: Application to interleukin 1. beta. United States: N. p., 1991. Web. doi:10.1021/bi00215a002.
Clore, G.M., Kay, L.E., Bax, A., & Gronenborn, A.M. Four-dimensional sup 13 C/ sup 13 C-edited nuclear Overhauser Enhancement Spectroscopy of a protein in solution: Application to interleukin 1. beta. United States. doi:10.1021/bi00215a002.
Clore, G.M., Kay, L.E., Bax, A., and Gronenborn, A.M. Tue . "Four-dimensional sup 13 C/ sup 13 C-edited nuclear Overhauser Enhancement Spectroscopy of a protein in solution: Application to interleukin 1. beta". United States. doi:10.1021/bi00215a002.
@article{osti_5613529,
title = {Four-dimensional sup 13 C/ sup 13 C-edited nuclear Overhauser Enhancement Spectroscopy of a protein in solution: Application to interleukin 1. beta},
author = {Clore, G.M. and Kay, L.E. and Bax, A. and Gronenborn, A.M.},
abstractNote = {A four-dimensional {sup 13}C/{sup 13}C-edited NOESY experiment is described which dramatically improves the resolution of protein NMR spectra and enables the straightforward assignment of nuclear Overhauser effects involving aliphatic and/or aromatic protons in larger proteins. The experiment is demonstrated for uniformly (>95{percent}) {sup 13}C-labeled interleukin 1{beta}, a protein of 153 residues and 17.4 kDa, which plays a key role in the immune response. NOEs between aliphatic and/or aromatic protons are first spread out into a third dimension by the {sup 13}C chemical shift of the carbon atom attached to the originating proton and subsequently into a fourth dimension by the {sup 13}C chemical shift of the carbon atom attached to the destination proton. Thus, each NOE cross peak is labeled by four chemical shifts. By this means, ambiguities in the assignment of NOEs that arise from chemical shift overlap and degeneracy are completely removed. Further, NOEs between protons with the same chemical shifts can readily be detected providing their attached carbon atoms have different {sup 13}C chemical shifts. The design of the pulse sequence requires special care to minimize the level of artifacts arising from undesired coherence transfer pathways, and in particular those associated with diagonal peaks which correspond to magnetization that has not been transferred from one proton to another. The 4D {sup 13}C/{sup 13}C-edited NOESY experiment is characterized by high sensitivity as the through-bond transfer steps involve the large {sup 1}J{sub CH} (130 Hz) couplings, and it is possible to obtain high-quality spectra on 1-2 mM samples of {sup 13}C-labeled protein in as little as 3 days. This experiment should open up the application of protein structure determination by NMR to a large number of medium-sized proteins (150-300 residues) of biological interest.},
doi = {10.1021/bi00215a002},
journal = {Biochemistry; (USA)},
issn = {0006-2960},
number = ,
volume = 30:1,
place = {United States},
year = {1991},
month = {1}
}