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KMID : 1059520230670020081
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Journal of the Korean Chemical Society
2023 Volume.67 No. 2 p.81 ~ p.88
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Study of HubWA Protein Folding Reaction by Measuring the Stability of Folding Intermediate
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Park Soon-Ho
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Abstract
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The contribution of hydrophobic residues to the protein folding reaction was studied by using HubWA variant proteins with I and L to V mutation. Folding kinetics of all V variant proteins was observed to be satisfied by a three-state on- pathway mechanism, U ? I ? N, where U, I, and N represent unfolded, intermediate, and native state, respectively. Three- state folding reaction was quantitatively analyzed and the free energy of folding of each elementary reactions and overall fold- ing reaction, ΔGoUI, ΔGoIN, and ΔGoUN, were obtained. From the ratio of free energy difference between the variant protein and HubWA, ΔΔGoUI/ΔΔGoUN (ΔΔGoUI = ΔGoUI (variant protein) ? ΔGoUI (HubWA) and ΔΔGoUN = ΔGoUN (variant protein) ? ΔGoUN (HubWA)), the contribution of hydrophobic residues to HubWA folding was analyzed. The residues which are located in the hydrophobic core between α-helix and β-sheet, I3, I13, L15, I30, L43, I61 and L67, showed ΔΔGoUI/ΔΔGoUN value of ~0.5 when each of these residues was mutated to V, indicating that these residues form relatively solid hydrophobic core in the intermediate state. Residues located at the end of secondary structures and loop, I23, L69 and I36 showed ΔΔGoUI/ΔΔGoUN value below 0.4 when each of these residues was mutated to V, indicating that the region containing these residues are loosely formed in the intermediate state. V17A, L50V and L56V showed fairly high ΔΔGoUI/ΔΔGoUN value of ~0.8. Since L50 and L56 are located in the region containing long loop (residue 46 to 62), it is suggested that the high ΔΔGoUI/ΔΔGoUN value of these residues prevents the formation of aggregate at the early stage of folding reaction.
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KEYWORD
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Hydrophobic interactions and protein folding, Protein folding intermediate
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