| Co-Investigator(Kenkyū-buntansha) |
KANAOKA Yoshihide Osaka Bioscience Institute, 2nd Dept., Researcher, 研究員 (40271514)
MATSUMURA Hitoshi Osaka Bioscience Institute, 2nd Dept., Vice-Head, 副部長 (50173886)
HAYAISHI Osamu Osaka Bioscience Institute, Director, 2nd Dept., Head, 所長, 部長 (40025507)
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| Research Abstract |
Prostaglandin (PG) D2 is a major prostanoid produced in the central nervous system of rats and humans. PGD2 is involved in a variety of central actions, i.e., it induces s sleep in rats and monkeys and regulates odor and pain responses. Lipocalin-type PGD synthase is responsible for biosynthesis of PGD2 in the central nervous system and is considered to be the key enzyme for regulation of physilogical sleep. We isolated the cDNA and gene for lipocalin-type PGD synthase and showed that it is the only enzyme among members of the lipocalin gene family which is composed of various secretory proteins. The enzyme was localized in the central nervous system and male genital organs and was secreted into the cerebrospinal fluid and seminal plasma as beta-trace. The site-directed mutagenesis and chemical modification studies revealed that Cys-65 of the enzyme is an essential amino acid residue for the catalytic reaction. The enzyme is, therefore, considered to have evolved from an non-enzymic ancestor protein by aquiring the active amino acid residue, Cys-65, for the catalytic reaction. However, we also found that the enzyme still maintained characteristics as a lipocalin, i.e., it bound specifically lipophilic substances, such as retinoids, thyroids and bile pigments, with affinities of the Kd values from 70 nM to 2 muM which were comparable with those of other hydrophobic ligand carrier proteins. We then generated the knockout mice lacking the gene for lipocalin-type PGD synthase and found the functional abnormality of the homozygous mice in terms of their reproductive rate and pain responses. We also crystallized the recombinant mouse lipocalin-type PGD synthase and obtained the diffraction data by X-ray crystallographic analysis.
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