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Partial NMR assignments and secondary structure mapping of the isolated α subunit of Escherichia coli tryptophan synthase, a 29‐kD TIM barrel protein
Author(s) -
Vadrevu Ramakrishna,
Falzone Christopher J.,
Matthews C. Robert
Publication year - 2003
Publication title -
protein science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.353
H-Index - 175
eISSN - 1469-896X
pISSN - 0961-8368
DOI - 10.1110/ps.0221103
Subject(s) - triosephosphate isomerase , chemistry , heteronuclear molecule , heteronuclear single quantum coherence spectroscopy , protein secondary structure , barrel (horology) , escherichia coli , tryptophan synthase , protein subunit , nuclear magnetic resonance spectroscopy , atp synthase , stereochemistry , chemical shift , crystallography , protein folding , hydrogen–deuterium exchange , biochemistry , enzyme , mass spectrometry , materials science , chromatography , composite material , gene
Abstract The α subunit of tryptophan synthase (αTS) from S. typhimurium belongs to the triosephosphate isomerase (TIM) or the (β/α) 8 barrel fold, one of the most common structures in biology. To test the conservation of the global fold in the isolated Escherichia coli homolog, we have obtained a majority of the backbone assignments for the 29‐kD αTS by using standard heteronuclear multidimensional NMR methods on uniformly 15 N‐ and 15 N/ 13 C‐labeled protein and on protein selectively 15 N‐labeled at key hydrophobic residues. The secondary structure mapped by chemical shift index, nuclear Overhauser enhancements (NOEs), and hydrogen‐deuterium (H‐D) exchange, and several abnormal chemical shifts are consistent with the conservation of the global TIM barrel fold of the isolated E. coli αTS. Because most of the amide protons that are slow to exchange with solvent correspond to the β‐sheet residues, the β‐barrel is likely to play an important role in stabilizing the previously detected folding intermediates for E. coli αTS. A similar combination of uniform and selective labeling can be extended to other TIM barrel proteins to obtain insight into the role of the motif in stabilizing what appear to be common partially folded forms.