Molecular characterization of a novel type of prostamide/prostaglandin F synthase, belonging to the thioredoxin-like superfamily.
Prostaglandin F (PGF) ethanolamide (prostamide F) synthase, which catalyzed the reduction of prostamide H(2) to prostamide F(2alpha), was found in mouse and swine brain. The enzyme was purified from swine brain, and its amino acid sequence was defined. The mouse enzyme consisted of a 603-bp open reading frame coding for a 201-amino acid polypeptide with a molecular weight of 21,669. The amino acid sequence placed the enzyme in the thioredoxin-like superfamily with Cys(44) being the active site. The enzyme expressed in Escherichia coli as well as the native enzyme catalyzed not only the reduction of prostamide H(2) to prostamide F(2alpha) but also that of PGH(2) to PGF(2alpha). The V(max) and K(m) values for prostamide H(2) were about 0.25 micromol/min.mg of protein and 7.6 microm, respectively, and those for PGH(2) were about 0.69 micromol/min.mg of protein and 6.9 microm, respectively. Neither PGE(2) nor PGD(2) served as a substrate for this synthase. Based on these data, we named the enzyme prostamide/PGF synthase. Although the enzyme showed a broad specificity for reductants, reduced thioredoxin preferentially served as a reducing equivalent donor for this enzyme. Moreover, Northern and Western blot analyses in addition to the prostamide F synthase activity showed that the enzyme was mainly distributed in the brain and spinal cord, and the immunohistochemical study in the spinal cord showed that the enzyme was found mainly in the cytosol. These results suggest that prostamide/PGF synthase may play an important functional role in the central nervous system.
Moriuchi
Hiroshi
H
Division of Life Science, Graduate School of Integrated Science and Art, University of East Asia, 2-1 Ichinomiyagakuen, Shimonoseki, Yamaguchi 751-8503, Japan.
Koda
Noriko
N
Okuda-Ashitaka
Emiko
E
Daiyasu
Hiromi
H
Ogasawara
Kensuke
K
Toh
Hiroyuki
H
Ito
Seiji
S
Woodward
David F
DF
Watanabe
Kikuko
K
D000595:Amino Acid Sequence; D000818:Animals; D001921:Brain / Q000201:enzymology; D017124:Conserved Sequence; D003600:Cytosol / Q000201:enzymology; D015232:Dinoprostone / Q000031:analogs & derivatives / Q000378:metabolism; D005260:Female; D006801:Humans; D006911:Hydroxyprostaglandin Dehydrogenases / Q000145:classification / Q000235:genetics* / Q000378:metabolism; D007700:Kinetics; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D013045:Species Specificity; D013379:Substrate Specificity; D013552:Swine; D013879:Thioredoxins / Q000145:classification / Q000235:genetics* / Q000378:metabolism
C507368:prostamide H2; D013879:Thioredoxins; D006911:Hydroxyprostaglandin Dehydrogenases; C031051:prostaglandin-F synthase; D015232:Dinoprostone