OneG-Vali: a computational tool for detecting, estimating and validating cryptic intermediates of proteins under native conditions

Abstract

Understanding structural excursions of proteins under native conditions at residue level resolutions is crucial to map energy landscapes of proteins and also to solve the ‘Levinthal paradox’ of protein folding. Native-state hydrogen–deuterium (NS H/D) exchange methods are powerful to structurally characterize cryptic intermediates (CIs) populating sparsely in the unfolding kinetics of proteins under conditions favoring folded conformations of the proteins. However, the methods are not applicable to proteins that are susceptible to denaturation or degradation or aggregation in the course of exchange experiments and also to proteins that are smaller in size (<10 kDa) in general. We have herein demonstrated a novel computational tool, OneG-Vali, which predicts the possible existence of cryptic intermediates of proteins in a qualitative and quantitative manner. And, the tool validates the prediction efficiency by comparing multistate unfolding curves defined by the tool with pseudo two-state unfolding curves of the proteins determined by macroscopic methods. In addition, the OneG-Vali facilitates accounting for the effect of cis–trans proline isomerization on estimating the population of CIs defined by the tool and also by experimental methods. The prediction accuracy of the tool is validated using proteins such as cytochrome c, apocytochrome b₅₆₂, the third domain of PDZ and T4 lysozyme. The OneG-Vali is implemented using CGI-Perl and it can be freely accessed and instantly used at http://sblab.sastra.edu/oneg-vali.html.