1997 年度実績報告書

ジーンターゲテイングによる分泌型プロスタグランジンD合成酵素の機能解析

研究課題

研究課題/領域番号	07457033
研究機関	(財)大阪バイオサイエンス研究所
研究代表者	裏出良博 (財)大阪バイオサイエンス研究所, 第2研究部, 副部長 (10201360)
研究分担者	金岡禧秀 (財)大阪バイオサイエンス研究所, 第2研究部, 研究員 (40271514) 松村人志 (財)大阪バイオサイエンス研究所, 第2研究部, 副部長 (50173886) 早石修 (財)大阪バイオサイエンス研究所, 第2研究部, 所長、部長 (40025507)
キーワード	睡眠 / 覚醒 / プロスタグランジンD2 / プロスタグランジンD合成酵素 / 遺伝子工学 / ノックアウトマウス / リポカリン / ベータートレース
研究概要	ラットおよびヒト脳における主要プロスタグランジン(PG)であるPGD2は、強力な睡眠誘発作用を示し、痛覚や臭覚の制御などの多彩な中枢作用を示す。中枢神経系におけるPGD2の生合成反応を触媒し生理的睡眠調節の鍵を握る酵素と考えられているリポカリン型PGD合成酵素は、そのcDNAクローニングの結果、各種の分泌蛋白質で構成されるリポカリンファミリーに属する唯一の酵素であることが判明した。本酵素は脳・脊髄などの中枢神経系や雄性生殖器に分布し、bトレース蛋白質として脳脊髄液や精液中に分泌される。本酵素は疎水性リガンドの輸送蛋白質が進化の過程で活性中心アミノ酸(Cys65)を獲得して酵素に進化したと考えられるが、他のリポカリン蛋白質と同様、レチノイドや甲状腺ホルモン・胆汁色素などの疎水性物質を特異的に結合する特性を保持していた。そこで、マウスの本酵素遺伝子を単離し、本酵素遺伝子を欠損したノックアウトマウスを作製した。その結果、遺伝子欠損ホモ・マウスが得られ、本酵素の遺伝子欠損は致死性を伴わないことが判明した。又、得られた遺伝子欠損マウスを用いて、その遺伝子欠損が繁殖率の異常や痛覚反応の異常をもたらすことを証明した。さらに、マウスの本酵素cDNAを大腸菌内で発現させた遺伝子組換え型酵素を用いて、世界で初めてリポカリン型PGD合成酵素の結晶化に成功した。

研究成果
(24件)

すべてその他

すべて文献書誌 (24件)

[文献書誌] Yoshihiro Urada: "Lipid Mediators Cell Signalling" J.Neurosci.257-273. 12. 257-273 (1995)
[文献書誌] Makoto Murakami: "Interleukin-4 suppresses c-Kit ligand-induced expression of cytosolic phospholipase A2 and prostaglandin endoperoxide synthase-2,and their roles in separate pathways of eicosanoid synthesis in mouse bone marrow-derived mast sells." Proc.Natl.Acad.Sci.USA. 92. 6107-6111 (1995)
[文献書誌] Bettina Blodorn: "he major site of mRNA expression for the b-trace protein (prostaglandin D synthase) in human brain." Neurosci.Lett.209. 117-120 (1996)
[文献書誌] Hiroshi Oda: "Quantitative sandwich immunosorbent assay of human secretory prostaglandin D synthase (b-trace)." Proc.Japan Acad.72. 108-111 (1996)
[文献書誌] Carsten T.Beuckmann: "Lipocalin-type prostaglandin D synthase (b-trace) is located in pigmented epithelial cells of rat retina and accumulates within interphotoreceptor matrix." J.Neurosci.16. 6119-6124 (1996)
[文献書誌] Yoshihiro Urade: "Molecular mechanism of sleep regulation by prostaglandin D2. J." Lipid Mediators Cell Signalling. 14. 71-82 (1996)
[文献書誌] Dimitrios N.Melegos: "Immunofluorometric assay of prostaglandin D synthase (b-trace) and preliminary clinical applications." Clin.Chem.42. 1984-1991 (1996)
[文献書誌] Hiroyuki Toh: "IGlutathione-independent prostaglandin D synthase as a lead molecule for designing new functional proteins." Protein Engineering. 9. 1067-1082 (1996)
[文献書誌] Tetsumori Yamashima: "IProstaglandin D synthase (b-trace) in human arachnoid and meningioma cells : roles as a cell marker or in CSF absorption,tumorigenesis,and calcification process." J.Neurosci. 17. 2376-2382 (1997)
[文献書誌] Toshiki Tanaka: "Lipocalin-type prostaglandin D synthase (b-trace) in a newly recognized type of retinoid transporter." J.Biol.Chem.272. 15789-15795 (1997)
[文献書誌] Luis F.Garcia-Fernandez: "Hypothyroidism alters the expression of prostaglandin D2 synthase/β-trace in specific areas of the developing rat brain. Eur." J.Neurosci.9. 1566-1573 (1997)
[文献書誌] Atsushi Hiraoka: "Analysis of low-molecular-mass proteins in cerebrospinal fluid by sodium dodecyl sulfate capillary electrophoresis." J.Chromatogr.697. 141-147 (1997)
[文献書誌] Peter Dieter: "Arachidonic acid cascade in kupffer cells." Cells Hepatic Sinusoid. 6. 355-356 (1997)
[文献書誌] Yoshihide Kanaoka: "Cloning and crystal structure of hematopoietic prostaglandin D synthase." Cell. 90. 1085-1095 (1997)
[文献書誌] Ishtiaq Mahmud: "Prostaglandin D synthase in human megakaryoblastic cells." J.Biol.Chem.272. 28263-28266 (1997)
[文献書誌] Yutaka Eguchi: "Expression of lipocalin-type prostaglandin D synthase (b-trace) in human heart and its accumulation in the coronary circulation of angina patients." Proc.Natl.Acad.Sci.USA.94. 14689-14694 (1997)
[文献書誌] Yoshihiro Tokugawa: "Lipocalin-type prostaglandin D synthase in human male reproductive organs and seminal plasma." Biol.Reprod. 58. 600-607 (1998)
[文献書誌] Yoshihiro Urade: "Gene engineering studies on sleep" Jpn.Clin.Med.56. 488-492 (1998)
[文献書誌] Yoshihiro Urada: "Gene engineering studies on sleep" Jpn.Clin.Med.56. 488-492 (1998)
[文献書誌] Atsushi Hiraoka: "Sodium dodecyl sulfate capillary electrophoretic analysis of molecular mass heterogeneity of b-trace protein in cerebrospinal fluid from patients with central nervous system diseases." J.Chromatogr.(in press). (1998)
[文献書誌] Dmitry Gerashchenko: "Localization of lipocalin-type prostanoid D synthase (b-trace) in iris,cilliary body,and eye fluids." Invest.Ophthalmol.Vis.Sci.(in press). (1998)
[文献書誌] Robyn L.Gerena: "Identification of a fertility-associated protein in bull seminal plasma as lipocalin-type prostaglandin D synthase." Biol.Reprod.(in press). (1998)
[文献書誌] Luis F.Garcia-Fernandez: "Identification of a thyroid hormone response element in the 5'-flanking region of the rat lipocalin-type prostaglandin D2 synthase (b-trace) gene." Mol.Brain.Res.(in press). (1998)
[文献書誌] Dmitry Gerashchenko: "Dominant expression of rat prostanoid DP receptor mRNA in leptomeninges,inner segments of photoreceptor cells,iris epithelium,and cilliary processes." J.Neurochem. (in press). (1998)

1997 年度 実績報告書

ジーンターゲテイングによる分泌型プロスタグランジンD合成酵素の機能解析

研究代表者

裏出 良博 (財)大阪バイオサイエンス研究所, 第2研究部, 副部長 (10201360)

研究成果

[文献書誌] Yoshihiro Urada: "Lipid Mediators Cell Signalling" J.Neurosci.257-273. 12. 257-273 (1995)

[文献書誌] Bettina Blodorn: "he major site of mRNA expression for the b-trace protein (prostaglandin D synthase) in human brain." Neurosci.Lett.209. 117-120 (1996)

[文献書誌] Hiroshi Oda: "Quantitative sandwich immunosorbent assay of human secretory prostaglandin D synthase (b-trace)." Proc.Japan Acad.72. 108-111 (1996)

[文献書誌] Carsten T.Beuckmann: "Lipocalin-type prostaglandin D synthase (b-trace) is located in pigmented epithelial cells of rat retina and accumulates within interphotoreceptor matrix." J.Neurosci.16. 6119-6124 (1996)

[文献書誌] Yoshihiro Urade: "Molecular mechanism of sleep regulation by prostaglandin D2. J." Lipid Mediators Cell Signalling. 14. 71-82 (1996)

[文献書誌] Dimitrios N.Melegos: "Immunofluorometric assay of prostaglandin D synthase (b-trace) and preliminary clinical applications." Clin.Chem.42. 1984-1991 (1996)

[文献書誌] Hiroyuki Toh: "IGlutathione-independent prostaglandin D synthase as a lead molecule for designing new functional proteins." Protein Engineering. 9. 1067-1082 (1996)

[文献書誌] Tetsumori Yamashima: "IProstaglandin D synthase (b-trace) in human arachnoid and meningioma cells : roles as a cell marker or in CSF absorption,tumorigenesis,and calcification process." J.Neurosci. 17. 2376-2382 (1997)

[文献書誌] Toshiki Tanaka: "Lipocalin-type prostaglandin D synthase (b-trace) in a newly recognized type of retinoid transporter." J.Biol.Chem.272. 15789-15795 (1997)

[文献書誌] Luis F.Garcia-Fernandez: "Hypothyroidism alters the expression of prostaglandin D2 synthase/β-trace in specific areas of the developing rat brain. Eur." J.Neurosci.9. 1566-1573 (1997)

[文献書誌] Atsushi Hiraoka: "Analysis of low-molecular-mass proteins in cerebrospinal fluid by sodium dodecyl sulfate capillary electrophoresis." J.Chromatogr.697. 141-147 (1997)

[文献書誌] Peter Dieter: "Arachidonic acid cascade in kupffer cells." Cells Hepatic Sinusoid. 6. 355-356 (1997)

[文献書誌] Yoshihide Kanaoka: "Cloning and crystal structure of hematopoietic prostaglandin D synthase." Cell. 90. 1085-1095 (1997)

[文献書誌] Ishtiaq Mahmud: "Prostaglandin D synthase in human megakaryoblastic cells." J.Biol.Chem.272. 28263-28266 (1997)

[文献書誌] Yutaka Eguchi: "Expression of lipocalin-type prostaglandin D synthase (b-trace) in human heart and its accumulation in the coronary circulation of angina patients." Proc.Natl.Acad.Sci.USA.94. 14689-14694 (1997)

[文献書誌] Yoshihiro Tokugawa: "Lipocalin-type prostaglandin D synthase in human male reproductive organs and seminal plasma." Biol.Reprod. 58. 600-607 (1998)

[文献書誌] Yoshihiro Urade: "Gene engineering studies on sleep" Jpn.Clin.Med.56. 488-492 (1998)

[文献書誌] Yoshihiro Urada: "Gene engineering studies on sleep" Jpn.Clin.Med.56. 488-492 (1998)

[文献書誌] Atsushi Hiraoka: "Sodium dodecyl sulfate capillary electrophoretic analysis of molecular mass heterogeneity of b-trace protein in cerebrospinal fluid from patients with central nervous system diseases." J.Chromatogr.(in press). (1998)

[文献書誌] Dmitry Gerashchenko: "Localization of lipocalin-type prostanoid D synthase (b-trace) in iris,cilliary body,and eye fluids." Invest.Ophthalmol.Vis.Sci.(in press). (1998)

[文献書誌] Robyn L.Gerena: "Identification of a fertility-associated protein in bull seminal plasma as lipocalin-type prostaglandin D synthase." Biol.Reprod.(in press). (1998)

[文献書誌] Luis F.Garcia-Fernandez: "Identification of a thyroid hormone response element in the 5'-flanking region of the rat lipocalin-type prostaglandin D2 synthase (b-trace) gene." Mol.Brain.Res.(in press). (1998)

[文献書誌] Dmitry Gerashchenko: "Dominant expression of rat prostanoid DP receptor mRNA in leptomeninges,inner segments of photoreceptor cells,iris epithelium,and cilliary processes." J.Neurochem. (in press). (1998)

1997 年度実績報告書

裏出良博 (財)大阪バイオサイエンス研究所, 第2研究部, 副部長 (10201360)