Immunochemical identification of the pssA gene product as phosphatidylserine synthase I of Chinese hamster ovary cells.
We have previously shown that a Chinese hamster ovary (CHO) cell mutant defective in phosphatidylserine synthase I recovers the enzyme activity on transfection with a pssA cDNA clone isolated from the parental CHO-K1. The resultant transfectant, CDT-1, exhibited about 20-fold higher specific activity of the enzyme in the membrane fraction than CHO-K1 cells. Polyclonal antibodies against two peptides of the predicted pssA product cross-reacted with a membrane protein having an apparent molecular mass of 42 kDa, which was overproduced in CDT-1 cells. By immunoprecipitation with the antibody, phosphatidylserine synthase I activity as well as the 42-kDa protein was eliminated from solubilized membrane proteins of CDT-1 cells. Both the enzyme activity and the 42-kDa protein of CHO-K1 cells were enriched in the mitochondria-associated membrane fraction and the microsome fraction, but neither was enriched in the mitochondria fraction or the cytosol fraction. These results suggest that the pssA gene encodes phosphatidylserine synthase I.
The Department of Biochemistry and Cell Biology, National Institute of Health, Tokyo, Japan.
K
K
Saito
O
O
Kuge
Y
Y
Akamatsu
M
M
Nishijima
D000818:Animals; D000906:Antibodies; D016466:CHO Cells; D006224:Cricetinae; D003600:Cytosol / Q000201:enzymology; D018076:DNA, Complementary; D015151:Immunoblotting; D007700:Kinetics; D008565:Membrane Proteins / Q000032:analysis / Q000096:biosynthesis / Q000378:metabolism*; D008861:Microsomes / Q000201:enzymology; D008928:Mitochondria / Q000201:enzymology; D019884:Nitrogenous Group Transferases; D013347:Subcellular Fractions / Q000201:enzymology; D014162:Transfection; D014166:Transferases / Q000032:analysis / Q000096:biosynthesis / Q000378:metabolism*
D000906:Antibodies; D018076:DNA, Complementary; D008565:Membrane Proteins; D014166:Transferases; D019884:Nitrogenous Group Transferases; C022370:phospholipid serine base exchange enzyme