Charge-reversion mutagenesis of Dictyostelium actin to map the surface recognized by myosin during ATP-driven sliding motion

Masaaki Johara, Yoko Yano Toyoshima, Akihiko Ishijima, Hiroaki Kojima, Toshio Yanagida, Kazuo Sutoh

Research output: Contribution to journalArticlepeer-review

70 Citations (Scopus)

Abstract

Amino acid residues D24/D25, E99/E100, E360/E361, and D363/E364 in subdomain 1 of Dictyostelium actin were replaced with histidine residues by site-directed mutagenesis. Mutant actins were expressed in Dictyostelium cells and purified to homogeneity. The sliding movement of mutant actin filaments on heavy meromyosin attached to a glass surface was measured to assess the effect of the mutation on the motility of actin. For two C-terminal mutants, force generated by a single actin filament and myosin was also measured. These measurements indicated that both D24/D25 and E99/E100 are involved in ATP-driven sliding, whereas E360/E361/ D363/E364 are not essential for ATP-driven sliding and force generation.

Original languageEnglish
Pages (from-to)2127-2131
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume90
Issue number6
DOIs
Publication statusPublished - 1993 Mar 15

Keywords

  • Actin-activated myosin ATPase
  • Force generation
  • In vitro motility assay
  • Site-directed mutagenesis

ASJC Scopus subject areas

  • General

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