Xiaoyi Yao, Stephanie Grade, Willy Wriggers, and Peter A. Rubenstein. His73, Often Methylated, is an Important Structural Determinant for Actin - A Mutagenic Analysis of H73 of Yeast Actin. Journal of Biological Chemistry. Vol. 274, pp. 37443-37449, 1999.
His73 has been proposed to regulate the release of Pi from the interior of actin following polymerization-dependent hydrolysis of bound ATP. Although it is a 3-methylhistidine in the vast majority of actins, H73 is unmethylated in S. cerevisiae actin. We mutated H73 in yeast actin to R,K,A,Q, and E and detected no altered phenotypes associated with the mutations in vivo. However, they significantly affect actin function in vitro. Substitution of the more basic residues resulted in enhanced thermal stability, decreased rate of nucleotide exchange, and decreased susceptibility to controlled proteolysis relative to wild type actin. The opposite effects are observed with the neutral and anionic substitutions. All mutations reduced the rate of polymerization. Molecular dynamics simulations predict a new conformation for the H73 imidazole in the absence of a methyl group. It also predicts that R73 tightens and stabilizes the actin and that E73 causes a rearrangement of the bottom of actin's interdomain cleft leading possibly to our observed destabilization of actin. Considering the exterior location of H73, this work indicates a surprisingly important role for the residue as a major structural determinant of actin and provides a clue to the impact caused by methylation of H73.
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