Identification of a cysteine residue important for the ATPase activity of C. elegans fidgetin homologue.
Based on the amino acid alignment, Caenorhabditis elegans F32D1.1 was identified to be a homologue of the mammalian fidgetin. We produced and purified the F32D1.1 protein by using a baculovirus-expression system. F32D1.1 has an ATPase activity, which is sensitive to N-ethylmaleimide. Km and Vmax for the ATPase activity of F32D1.1 were estimated to be 0.44 mM and 225 nmol/mg/min, respectively. When the cysteine at the position of 368 was mutated to alanine, the ATPase activity was greatly decreased; Vmax was decreased to one-sixth, while Km remained similar. These results suggest that the unique position of cysteine 368, located immediately downstream of the Walker A motif, plays an important role in the ATP hydrolysis process of C. elegans F32D1.1 protein.
Yakushiji
Yasufumi
Y
Division of Molecular Cell Biology, Institute of Molecular Embryology and Genetics, Kumamoto University, Kumamoto 862-0976, Japan.
Yamanaka
Kunitoshi
K
Ogura
Teru
T
D000251:Adenosine Triphosphatases / Q000037:antagonists & inhibitors / Q000145:classification / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D017173:Caenorhabditis elegans / Q000201:enzymology*; D029742:Caenorhabditis elegans Proteins / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D003545:Cysteine / Q000378:metabolism*; D004252:DNA Mutational Analysis; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D010802:Phylogeny; D016415:Sequence Alignment
D029742:Caenorhabditis elegans Proteins; D000251:Adenosine Triphosphatases; C496225:figl-1 protein, C elegans; D003545:Cysteine