Immunophotoaffinity labeling of binders of 1-methyladenine, the oocyte maturation-inducing hormone of starfish.
Starfish oocytes are arrested at the prophase stage of the first meiotic division in the ovary and resume meiosis by the stimulus of 1-methyladenine (1-MeAde), the oocyte maturation-inducing hormone of starfish. Putative 1-MeAde receptors on the oocyte surface have been suggested, but not yet been biochemically characterized. Immunophotoaffinity labeling, i.e., photoaffinity labeling combined with immunochemical detection, was attempted to detect unknown 1-MeAde binders including putative maturation-inducing hormone receptors in starfish oocytes. When the oocyte crude membrane fraction or its Triton X-100/EDTA extract was incubated with N6-[6-(5-azido-2-nitrobenzoyl)aminohexyl]carboxamidomethyl-1-methyladenine and then photo-irradiated, followed by western blotting with antibody that was raised against a 1-MeAde hapten, a single band with Mrof 47.5 K was detected. The band was lost when extract was heated at 100 °C. A similar 47.5 K band was detected in the crude membrane fraction of testis as well. Upon labeling with whole cells, this band was detected in immature and maturing oocytes, but only faintly in mature oocytes. As judged from these results, this 1-MeAde binder might be a possible candidate of the starfish maturation-inducing hormone receptors.
Department of Bioscience and Biotechnology, Graduate School of Natural Science and Technology, Okayama University, Tsushima-naka, Kita-ku, Okayama 700-8530, Japan. Electronic address: toraya@cc.okayama-u.ac.jp.
Tetsuo
T
Toraya
Department of Bioscience and Biotechnology, Graduate School of Natural Science and Technology, Okayama University, Tsushima-naka, Kita-ku, Okayama 700-8530, Japan.
Tetsuo
T
Kida
Department of Bioscience and Biotechnology, Graduate School of Natural Science and Technology, Okayama University, Tsushima-naka, Kita-ku, Okayama 700-8530, Japan.
Atsushi
A
Kuyama
Department of Bioscience and Biotechnology, Graduate School of Natural Science and Technology, Okayama University, Tsushima-naka, Kita-ku, Okayama 700-8530, Japan.
Shinjiro
S
Matsuda
Department of Bioscience and Biotechnology, Graduate School of Natural Science and Technology, Okayama University, Tsushima-naka, Kita-ku, Okayama 700-8530, Japan.
Seiichi
S
Tanaka
Department of Bioscience and Biotechnology, Graduate School of Natural Science and Technology, Okayama University, Tsushima-naka, Kita-ku, Okayama 700-8530, Japan.
Yo
Y
Komatsu
Department of Bioscience and Biotechnology, Graduate School of Natural Science and Technology, Okayama University, Tsushima-naka, Kita-ku, Okayama 700-8530, Japan.
Taro
T
Tsurukai
D000225:Adenine / Q000031:analogs & derivatives* / Q000378:metabolism; D000818:Animals; D015153:Blotting, Western / Q000379:methods; D005260:Female; D009865:Oocytes / Q000166:cytology / Q000378:metabolism*; D009866:Oogenesis; D019920:Photoaffinity Labels / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism*; D013215:Starfish / Q000502:physiology*
D019920:Photoaffinity Labels; C008407:1-methyladenine; D000225:Adenine
1-Methyladenine; Immunophotoaffinity labeling; Oocyte maturation; Receptor; Starfish