Molecular characterization of a novel type of prostamide/prostaglandin F synthase, belonging to the thioredoxin-like superfamily

Abstract

Prostaglandin F (PGF) ethanolamide (prostamide F) synthase, which catalyzed the reduction of prostamide H2 to prostamide F2α, 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 Cys44 being the active site. The enzyme expressed in Escherichia coli as well as the native enzyme catalyzed not only the reduction of prostamide H2 to prostamide F 2α but also that of PGH2 to PGF2α. The Vmax and Km values for prostamide H2 were about 0.25 μmol/min·mg of protein and 7.6 μM, respectively, and those for PGH2 were about 0.69 μmol/min·mg of protein and 6.9 μM, respectively. Neither PGE2 nor PGD2 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. © 2008 by The American Society for Biochemistry and Molecular Biology, Inc.