Dynamic structure of a highly ordered beta-sheet molten globule: multiple conformations with a stable core.

Elisar Barbar, George Barany, Clare Woodward

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Abstract

The structure of [14-38]Abu, a variant of bovine pancreatic trypsin inhibitor (BPTI) with only the 14-38 disulfide bridge intact, has been analyzed by two-dimensional 1H and 1H- 15N NMR. Except for the 18-24, 29-35 antiparallel β-sheet, residues in all regions of the molecule give two exchange cross peaks for each 1H; for one residue, Gly 37, three exchange cross peaks are observed. The presence of exchange cross peaks indicates that the residues sample conformations that interconvert on a time scale of milliseconds or longer. Over 90% of the NMR spectra have been assigned, including backbone and side chain atoms and their exchange cross peaks. Analyses of chemical shifts, chemical exchange, hydrogen isotope exchange, and NOEs indicate that [14-38]Abu at pH 4.5 and 1°C is an ensemble of interconverting conformations, in all of which the 18-24, 29-35 antiparallel β-sheet is native-like and intact. Outside the antiparallel β-sheet, residues undergo local order/disorder transitions. The stable structure of [14-38]Abu is not in the vicinity of the 14-38 disulfide bond but rather is in the slow-exchange core. NOE analysis indicates that the main tertiary interactions involve hydrophobic contacts with the rings of Tyr 21, Tyr 23, and Tyr 35. As a model for early folding intermediates, the structure of [14-38]Abu suggests that BPTI folding is initiated by stabilization of a turn existing in the unfolded protein and involves both local and nonlocal hydrophobic interactions.

Original languageEnglish (US)
Pages (from-to)11423-11434
Number of pages12
JournalBiochemistry
Volume34
Issue number36
DOIs
StatePublished - Sep 12 1995

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