Journal Article

Characterization of a Major Secretory Protein in the Cane Toad (Bufo marinus) Choroid Plexus as an Amphibian Lipocalin-type Prostaglandin D Synthase

Daisuke Irikura, Takashi Inui, Carsten T. Beuckmann, Kousuke Aritake, Gerhard Schreiber, Masashi Miyano, Tsuyoshi Inoue and Yoshihiro Urade

in The Journal of Biochemistry

Published on behalf of The Japanese Biochemical Society

Volume 141, issue 2, pages 173-180
Published in print February 2007 | ISSN: 0021-924X
Published online December 2006 | e-ISSN: 1756-2651 | DOI: https://dx.doi.org/10.1093/jb/mvm016
Characterization of a Major Secretory Protein in the Cane Toad (Bufo marinus) Choroid Plexus as an Amphibian Lipocalin-type Prostaglandin D Synthase

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Here we report the enzymatic and ligand-binding properties of a major secretory protein in the choroid plexus of cane toad, Bufo marinus, whose protein is homologous with lipocalin-type prostaglandin (PG) D synthase (L-PGDS) and is recombinantly expressed in Xenopus A6 cells and Escherichia coli. The toad protein bound all-trans retinal, bile pigment, and thyroid hormones with high affinities (Kd = 0.17 to 2.00 μM). The toad protein also catalysed the L-PGDS activity, which was accelerated in the presence of GSH or DTT, similar to the mammalian enzyme. The Km value for PGH2 (17 μM) of the toad protein was almost the same as that of rat L-PGDS (14 μM), whereas the turnover number (6 min−1) was approximately 28 fold lower than that of rat L-PGDS. Site-directed mutagenesis based on a modeled structure of the toad protein revealed that Cys59 and Thr61 residues were crucial for the PGDS activity. The quadruple Gly39Ser/Ala75Ser/Ser140Thr/Phe142Tyr mutant of the toad protein, resembling mouse L-PGDS, showed a 1.6 fold increase in the turnover number and a shift in the optimum pH for the PGDS activity from 9.0 to 8.5. Our results suggest that the toad protein is a prototype of L-PGDS with a highly functional ligand-binding pocket and yet with a primitive catalytic pocket.

Keywords: amphibian; lipocalin; lipocalin-type prostaglandin D synthase; prostaglandin D2; site-directed mutagenesis

Journal Article.  4586 words.  Illustrated.

Subjects: Biochemistry

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