Untitled

Abstract

Hydrogen exchange kinetics were measured on the native states of wild type staphylococcal nuclease and four mutants with values of mGuHCl (defined as dDeltaG/d[guanidine hydrochloride]) ranging from 0.8 to 1.4 of the wild type value. Residues within the five-strand beta-barrel of wild type and E75A and D77A, two mutants with reduced values of m GuHCl, were significantly more protected from exchange than expected on the basis of global stability as measured by fluorescence. In contrast, mutants V23A and M26G with elevated values of mGuHCl approach a flat profile of more or less constant protection independent of position in the structure. Differences in exchange protection between the C-terminus and the beta-barrel region correlate with mGuHCl, suggesting that a residual barrel-like structure becomes more highly populated in the denatured states of m- mutants and less populated in m+ mutants. Variations in the population of such a molten globule-like structure would account for the large changes in solvent accessible surface area of the denatured state thought to underlie m value effects.

Energetics of side-chain partitioning of β-signal residues in unassisted folding of a transmembrane β-barrel protein

“…This could arise from a change in the structure of PagP upon folding or the presence of residual structure in the unfolded state. Point mutations can alter the residual structure in the unfolded protein state without affecting the folded state and thereby change the m value (56). In such a case, mutants with residual structure in the unfolded state have a lower m value, whereas the mutants without any residual structure have a higher m value (37, 56).…”

“…Point mutations can alter the residual structure in the unfolded protein state without affecting the folded state and thereby change the m value (56). In such a case, mutants with residual structure in the unfolded state have a lower m value, whereas the mutants without any residual structure have a higher m value (37, 56). A comparison of the emission profiles of the unfolded PagP mutant proteins (<λ U >) suggests that the unfolded protein state might not possess any significant residual structure (supplemental Fig.…”

The fluorescence detected guanidine hydrochloride equilibrium denaturation of wild-type staphylococcal nuclease does not fit a three-state unfolding model

“…The first denatured state, D 1 , may be thought of as a folding intermediate if preferred. Let us further assume that D1 and D 2 states are indistinguishable by the probe of structure used here, fluorescence of W140, which is at the end of the C-terminal helix proposed to unfold in the first transition of a putative three-state unfolding [22-24]. In this model, the remaining structure, primarily the main beta barrel, then breaks down cooperatively in the subsequent step.…”

Characterization of human frataxin missense variants in cancer tissues

“…The thermal stability of FXN wild type and variants was investigated by continuously monitoring the ellipticity changes at 222 nm between 20°C and 95°C ( Figure S1) and between 95°C and 20°C The changes in m values may indicate differences in the solventexposed surface area upon unfolding between the variants and the wild type: decrease in m values is usually referred to a decrease in the solvent-exposed surface area upon unfolding. This is frequently ascribed to an increase in the compactness of the residual structure in the nonnative state ensemble, rather than to an increase of the accessible surface area of the native state (Pradeep & Udgaonkar, 2004;Wrabl & Shortle, 1999). Table S2).…”