Journal article
Directly observed nitrogen-15 NMR spectra of uniformly enriched proteins
Biochemistry (Easton), Vol.26(8), pp.2202-2207
04/01/1987
DOI: 10.1021/bi00382a020
Abstract
The proteins cytochrome c2, cytochrome c’, and ribulosebisphosphate carboxylase/oxygenase from Rhodospirillum rubrum were enriched in 15N by growth of the organism on 15NH4C1. The proteins were purified to homogeneity and studied by 15N NMR. Longitudinal and transverse relaxation times as well as the nuclear Overhauser effects were determined for various groups of the proteins which vary in molecular weight from 13 000 to 114000. The values of these parameters for the amide resonances or for groups thought to be rigid were consistent with the molecular weights of the proteins. Relaxation times of the amino-terminal a-amino groups and the side chain nitrogen atoms of arginine and lysine were consistent with much more rapid motion. Nitrogen atoms having bound protons were generally found to be decoupled from the protons by chemical exchange. Demonstrable 1H-15N coupling was taken as an indication that exchange was hindered, either by hydrogen bonding interactions or by inaccessibility of the group to solvent. Histidine side chain nitrogen atoms, which experience a large chemical shift upon protonation/deprotonation, were often found to be broadened beyond detectability by chemical exchange and tautomerization. Strategies for improving sensitivity and for obtaining specific peak assignments are also discussed.
Details
- Title: Subtitle
- Directly observed nitrogen-15 NMR spectra of uniformly enriched proteins
- Creators
- Gary M SmithLiping P YuDavid J Domingues - University of California, Davis
- Resource Type
- Journal article
- Publication Details
- Biochemistry (Easton), Vol.26(8), pp.2202-2207
- Publisher
- American Chemical Society
- DOI
- 10.1021/bi00382a020
- ISSN
- 0006-2960
- eISSN
- 1520-4995
- Language
- English
- Date published
- 04/01/1987
- Academic Unit
- Biochemistry and Molecular Biology; Medicine Administration
- Record Identifier
- 9984627190702771
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