title: Regulation of yeast actin behavior by interaction of charged residues across the interdomain cleft.
abstract: His(73) participates in the regulation of the nucleotide binding cleft conformation in yeast actin. Earlier molecular dynamics studies suggested that Asp(184) interacts with His(73) thereby stabilizing a "closed-cleft" G-actin. However, beta-actin in the open-cleft state shows a closer interaction of His(73) with Asp(179) than with Asp(184). We have thus assessed the relative importance of Asp(184) and Asp(179) on yeast actin stability and function. Neutral substitutions at 184 or 179 alone had little adverse effect on the monomer and polymerization behavior of actin. Arg or His at 184 in H73E actin partially rescued the monomeric properties of H73E actin, as demonstrated by near-normal thermostability and wild-type (WT)-like protease digestion patterns. ATP exchange was still considerably faster than with WT-actin although slower than that of H73E alone. However, polymerization of H73E/D184R and H73E/D184H is worse than with H73E alone. Conversely, D179R rescued all monomeric properties of H73E to near WT values and largely restored polymerization rate and filament thermostability. These results and new simulations of G-actin in the "open" state underscore the importance of the His(73)-Asp(179) interaction and suggest that the open and not the closed state of yeast actin may be favored in the absence of the methyl group of His(73).
authors.affiliation: Department of Biochemistry, University of Iowa College of Medicine, Iowa City, Iowa 52242, USA.
authors.forename: Xiaoyi
authors.initials: X
authors.lastname: Yao
authors.forename: Vinh
authors.initials: V
authors.lastname: Nguyen
authors.forename: Willy
authors.initials: W
authors.lastname: Wriggers
authors.forename: Peter A
authors.initials: PA
authors.lastname: Rubenstein
mesh_terms: D000199:Actins / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism; D001224:Aspartic Acid / Q000737:chemistry / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005656:Fungal Proteins / Q000378:metabolism*; D006639:Histidine / Q000737:chemistry; D008762:Methylhistidines / Q000737:chemistry; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D013696:Temperature; D013997:Time Factors
chemical_list: D000199:Actins; D005656:Fungal Proteins; D008762:Methylhistidines; D001224:Aspartic Acid; D006639:Histidine; D000255:Adenosine Triphosphate