cDNA cloning and expression of intracellular platelet-activating factor (PAF) acetylhydrolase II. Its homology with plasma PAF acetylhydrolase.
Platelet-activating factor (PAF) acetylhydrolase, which inactivates PAF by removing the acetyl group at the sn-2 position, is widely distributed in plasma and tissues. We previously demonstrated that tissue cytosol contains at least two types of PAF acetylhydrolase, isoforms Ib and II, and that isoform Ib is a heterotrimer comprising 45-, 30-, and 29-kDa subunits, whereas isoform II is a 40-kDa monomer. In this study, we isolated cDNA clones of bovine and human PAF acetylhydrolase isoform II. From the longest open reading frame of the cloned cDNAs, both bovine and human PAF acetylhydrolases II are predicted to contain 392 amino acid residues and to exhibit 88% identity with each other at the amino acid level. Both enzymes contain a Gly-X-Ser-X-Gly motif that is characteristic of lipases and serine esterases. Expression of isoform II cDNA in COS7 cells resulted in a marked increase in PAF acetylhydrolase activity. An immunoblot study using an established monoclonal antibody against the bovine enzyme revealed that the recombinant protein exists in the membranous fraction as well as the soluble fraction. Isoform II is expressed most abundantly in the liver and kidney in cattle, but low levels were also observed in other tissues. The amino acid sequence deduced from the cDNA of isoform II had no homology with any subunit of isoform Ib. Interestingly, however, the amino acid sequence of isoform II showed 41% identity with that of plasma PAF acetylhydrolase. Combined with previous data demonstrating that isoform II shows similar substrate specificity to plasma PAF acetylhydrolase, these results indicate that tissue type isoform II and the plasma enzyme may share a common physiologic function.
Hattori
K
K
Department of Health Chemistry, Faculty of Pharmaceutical Sciences, The University of Tokyo, Hongo 7-3-1, Bunkyo-ku, Tokyo 113, Japan.
Adachi
H
H
Matsuzawa
A
A
Yamamoto
K
K
Tsujimoto
M
M
Aoki
J
J
Hattori
M
M
Arai
H
H
Inoue
K
K
D043203:1-Alkyl-2-acetylglycerophosphocholine Esterase; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D002417:Cattle; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D006801:Humans; D007527:Isoenzymes / Q000235:genetics*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D010741:Phospholipases A / Q000097:blood / Q000235:genetics*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
D018076:DNA, Complementary; D007527:Isoenzymes; D010446:Peptide Fragments; D010741:Phospholipases A; D043203:1-Alkyl-2-acetylglycerophosphocholine Esterase