期刊
JOURNAL OF MOLECULAR BIOLOGY
卷 320, 期 2, 页码 311-319出版社
ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD
DOI: 10.1016/S0022-2836(02)00449-7
关键词
high pressure NMR; apomyoglobin; local unfolding; molten globule; volume theorem
Pressure-induced reversible conformational changes of sperm whale apomyoglobin have been studied between 30 bar and 3000 bar on individual residue basis by utilizing H-1/N-15 hetero nuclear single-quantum coherence two-dimensional NMR spectroscopy at pH 6.0 and 35 degreesC. Apomyoglobin showed a series of pressure-dependent NMR spectra as a function of pressure, assignable to the native (N), intermediates (1), molten globule (MG) and unfolded (U) conformers. At 30 bar, the native fold (N) shows disorder only in the F helix. Between 500 bar and 1200 bar, a series of locally disordered conformers I are produced, in which local disorder occurs in the C helix, the CID loop, the G helix and part of the H helix. At 2000 bar, most cross-peaks exhibit severe line-broadening, suggesting the formation of a molten globule, but at 3000 bar all the cross-peaks reappear, showing that the molten globule turns into a well-hydrated, mobile unfolded conformation U. Since all the spectral changes were reversible with pressure, apomyoglobin is considered to exist as an equilibrium mixture of the N, 1, MG and U conformers at all pressures. MG is situated at 2.4 +/- (0.1) kcal/mol above N at 1 bar and the unfolding transition from the combined N-I state to MG is accompanied by a loss of partial molar volume by 75 +/- (3) ml/mol. On the basis of these observations, we postulate a theorem that the partial molar volume of a protein decreases in parallel with the loss of its conformational order. (C) 2002 Elsevier Science Ltd. All rights reserved.
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