Effect of Substitutions in Surface Amino Acid on Energy Profile of Apomyoglobin
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Studies on the process of spontaneous protein folding into a unique native state are an important issue of molecular biology. Apomyoglobin from the sperm whale is a convenient model for these studies in vitro. Here, we present the results of equilibrium and kinetic experiments carried out in a study on the folding and unfolding of eight mutant apomyoglobin forms of with hydrophobic amino acid substitutions on the protein surface. Calculated values of apparent constants of folding/unfolding rates, as well as the data on equilibrium conformational transitions in the urea concentration range of 0–6 M at 11°C are given. Based on the obtained information on the kinetic properties of the studied proteins, a Φ-value analysis of the transition state has been performed and values of urea concentrations corresponding to the midpoint of the transition from the native to intermediate state have been determined for the given forms of mutant apomyoglobin. It has been found that a significant increase in the stability of the native state can be achieved by a small number of amino acid substitutions on the protein surface. It has been shown that the substitution of only one amino acid residue exclusively affects the height of the energy barrier that separates different states of apomyoglobin.
Keywordsapomyoglobin protein folding tryptophan fluorescence circular dichroism stopped-flow experiments chevron plot
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- 22.Melnik T.N., Majorina M.A., Larina D.S., et al. 2014. Independent of their localization in protein the hydrophobic amino acid residues have no effect on the molten globule state of apomyoglobin and the disulfide bond on the surface of apomyoglobin stabilizes this intermediate state. PLOS ONE. 9, e98645.CrossRefPubMedPubMedCentralGoogle Scholar
- 23.Samatova E.N., Melnik B.S., Balobanov V.A., et al. 2010. Folding intermediate and folding nucleus for I→N and U→I→N transitions in apomyoglobin: Contributions by conserved and nonconserved residues. Biophys. J. }}98}}, 1694–1702.Google Scholar
- 27.Strub C., Alies C., Lougarre A., et al. 2004. Mutation of exposed hydrophobic amino acids to arginine to increase protein stability. Biochemistry. 5, 1471–2091.Google Scholar