Molecular cloning, functional expression, and selective regulation of ovine prostaglandin H synthase-2.
Structural characterization for ovine prostaglandin H synthase-1 (PGHS-1) is extensive, but the corresponding structure for the homologous ovine PGHS-2 isoform is undefined. Accordingly, we isolated a full-length (3.4 kb) ovine PGHS-2 cDNA from a primary-culture cell model (ovine tracheal epithelial cells) originally described as containing both PGHS isoforms. Analysis of ovine PGHS-2 cDNA sequence indicated conservation of critical amino acid residues, but differences in other hydrophilic regions allowed for the development of an anti-peptide antibody highly selective for PGHS-2. Enzymatic activities of the recombinant ovine PGHS isozymes indicated significant differences in response to aspirin-acetylation consistent with the characteristics of endogenous cellular PGHS activities under basal and serum-induced conditions. The results fully account for previous evidence of two distinct PGHS activities in cultured airway epithelial cells and provide for additional definition of PGHS structure-function relationships.
Zhang
V
V
Department of Medicine, Washington University School of Medicine, St. Louis, Missouri 63110, USA.
O'Sullivan
M
M
Hussain
H
H
Roswit
W T
WT
Holtzman
M J
MJ
D000595:Amino Acid Sequence; D000818:Animals; D001241:Aspirin / Q000494:pharmacology; D001483:Base Sequence; D019556:COS Cells; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D003001:Cloning, Molecular; D017931:DNA Primers; D004848:Epithelium / Q000201:enzymology; D015971:Gene Expression Regulation, Enzymologic; D007527:Isoenzymes / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011451:Prostaglandin-Endoperoxide Synthases / Q000096:biosynthesis* / Q000302:isolation & purification / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism; D012756:Sheep; D014132:Trachea / Q000201:enzymology; D014162:Transfection
D017931:DNA Primers; D007527:Isoenzymes; D011994:Recombinant Proteins; D011451:Prostaglandin-Endoperoxide Synthases; D001241:Aspirin