<b>Purification and Characterization of the 15-kDa Protein from the Sperm Flagella of Salmonid </b><b>Fishes </b>

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Author(s)

    • ITOH Atsuko
    • Misaki Marine Biological Station, Graduate School of Science, the University of Tokyo
    • INABA Kazuo
    • Asamushi Marine Biological Station, Tohoku University
    • OHTAKE Hideki
    • Department of Physiology, Dokkyo University School of Medicine
    • MORISAWA Masaaki
    • Misaki Marine Biological Station, Graduate School of Science, the University of Tokyo

Abstract

Cyclic AMP-dependent phosphorylation of tyrosine residues in a protein with a molecular mass of 15-kDa is thought to play a key role in the initiation of sperm motility in salmonid fishes (7). In this study, flagellar proteins were solubilized with 7 M urea and fractionated by sucrose densitygradient isoelectrie focusing. A band relevant to 15-kDa was detected in the fraction at pH 5 on electrophoresis and analyzed using phosphotyrosine antibody. Analysis of the band by reversed phase chromatography showed a single peak, suggesting that the 15-kDa protein was completely purified. Amino acid sequence of His—Ile-Phe at the N-terminus was detected by analysis using peptide sequencer, and the fourth base could not be determined. Further analysis of the molecular structure of the protein by LC—MS/MS showed that partial amino acid sequence of the 15-kDa protein from tryptic fragments were similar to parts of the amino acid sequence of tubulin. The protein was not detected in inununoblots using anti-tubulin polyclonal antibody, suggesting that the 15-kDa protein may be not debris of the tubulin but a novel protein with a common partial sequence of tubulin.

Journal

  • Biomedical Research

    Biomedical Research 24(4), 153-164, 2003

    Biomedical Research Press

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