The effect of increasing the stability of non-native interactions on the folding landscape of the bacterial immunity protein Im9 |
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Authors: | Morton Victoria L Friel Claire T Allen Lucy R Paci Emanuele Radford Sheena E |
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Institution: | Astbury Centre for Structural Molecular Biology, University of Leeds, Leeds, LS2 9JT, UK. |
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Abstract: | How stabilising non-native interactions influence protein folding energy landscapes is currently not well understood: such interactions could speed folding by reducing the conformational search to the native state, or could slow folding by increasing ruggedness. Here, we examine the influence of non-native interactions in the folding process of the bacterial immunity protein Im9, by exploiting our ability to manipulate the stability of the intermediate and rate-limiting transition state (TS) in the folding of this protein by minor alteration of its sequence or changes in solvent conditions. By analysing the properties of these species using Phi-value analysis, and exploration of the structural properties of the TS ensemble using molecular dynamics simulations, we demonstrate the importance of non-native interactions in immunity protein folding and demonstrate that the rate-limiting step involves partial reorganisation of these interactions as the TS ensemble is traversed. Moreover, we show that increasing the contribution to stability made by non-native interactions results in an increase in Phi-values of the TS ensemble without altering its structural properties or solvent-accessible surface area. The data suggest that the immunity proteins fold on multiple, but closely related, micropathways, resulting in a heterogeneous TS ensemble that responds subtly to mutation or changes in the solvent conditions. Thus, altering the relative strength of native and non-native interactions influences the search to the native state by restricting the pathways through the folding energy landscape. |
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Keywords: | TS transition state SIm9 Im9 containing the mutations V37L and V71I U unfolded state I intermediate state N native state βX the β-Tanford value for species X SASA the solvent accessible surface area |
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