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Folding and Unfolding of TIM Monomers and Dimers

Department of Chemistry, Oakland University, Rochester, Michigan

Correspondence: Address reprint requests to John M. Finke, Dept. of Chemistry, Oakland University, Rochester, MI 48309-4477. Tel.: 248-370-3088; E-mail: finke{at}oakland.edu.

Kinetic simulations of the folding and unfolding of triosephosphate isomerase (TIM) from yeast were conducted using a single monomer TIM polypeptide chain that folds as a monomer and two TIM chains that fold to the native dimer structure. The basic protein model used was a minimalist G model using the native structure to determine attractive energies in the protein chain. For each simulation type—monomer unfolding, monomer refolding, dimer unfolding, and dimer refolding—thirty simulations were conducted, successfully capturing each reaction in full. Analysis of the simulations demonstrates four main conclusions. First, all four simulation types have a similar "folding order", i.e., they have similar structures in intermediate stages of folding between the unfolded and folded state. Second, despite this similarity, different intermediate stages are more or less populated in the four different simulations, with 1), no intermediates populated in monomer unfolding; 2), two intermediates populated with ß₂–ß₄ and ß₁–ß₅ regions folded in monomer refolding; 3), two intermediates populated with ß₂–ß₃ and ß₂–ß₄ regions folded in dimer unfolding; and 4), two intermediates populated with ß₁–ß₅ and ß₁–ß₅ + ß₆ + ß₇ + ß₈ regions folded in dimer refolding. Third, simulations demonstrate that dimer binding and unbinding can occur early in the folding process before complete monomer-chain folding. Fourth, excellent agreement is found between the simulations and MPAX (misincorporation proton alkyl exchange) experiments. In total, this agreement demonstrates that the computational G model is accurate for TIM and that the energy landscape of TIM appears funneled to the native state.