Structure and stability of the molten globule state of guinea pig .alpha.-lactalbumin: A hydrogen exchange study

Citations Over TimeTop 1% of 1993 papers

Abstract

A partially folded state of guinea pig alpha-lactalbumin (the A-state or molten globule state), formed by denaturation at low pH, has been studied using hydrogen exchange methods. The overall distribution of exchange kinetics, measured by 1-D NMR, suggests that fewer than 20 amides in the structure are involved in highly persistent residual structure, although CD results suggest that many other parts of the chain are folded, for a significant proportion of the time, into less stable structural elements. The pH-jump experiments show that some amides that are strongly protected from exchange in the native state become freely accessible in the A-state but that conversely a majority, at least, of those that are slow to exchange in the A-state retain that protection in the native state. This suggests that the persistent structure in the A-state is native-like although the possibility that nonnative like structural elements persist cannot be eliminated. Resonance assignments for key residues in the NMR spectrum of the native state have enabled us to use the pH-jump method also to identify the majority of the most protected amides in the A-state: they are located in two hydrophobic segments, corresponding to the B- and C-helices of the native structure. This strongly suggests that the most persistent structure of the A-state includes these regions. A variety of lines of evidence, including fluorescence quenching data and, most remarkably, very effective protection from exchange of an indole NH in a tryptophan side chain, suggest that some form of hydrophobic core in the helical domain of the native structure persists in the A-state, although without the stereochemical rigidity of the native tertiary structure. The other domain of the native structure, including the beta-sheet, appears not to contain structural elements which persist to the same extent in the A-state, emphasizing that the molten globule is highly heterogeneous, in terms of the stability and specificity of both backbone and side chain interactions.

Related Papers

• → Different Subdomains are Most Protected From Hydrogen Exchange in the Molten Globule and Native States of Human α-Lactalbumin(1995)193 cited
• → Pathway of disulfide-coupled unfolding and refolding of bovine .alpha.-lactalbumin(1993)107 cited
• → Contribution of the 6-120 disulfide bond of .alpha.-lactalbumin to the stabilities of its native and molten globule states(1992)47 cited
• → The Native-like Tertiary Fold in Molten Globule α-Lactalbumin Appears to be Controlled by a Continuous Phase Transition(1996)33 cited
• → Local and long-range interactions in the molten globule state: a study of chimeric proteins of bovine and human α-lactalbumin(2000)33 cited