Testicular Angiotensin-converting enzyme with different glycan modification: characterization on glycosylphosphatidylinositol-anchored protein releasing and dipeptidase activities.
We have previously found that the angiotensin-converting enzyme (ACE) carries GPI-anchored protein releasing activity (GPIase) as well as dipeptidase activity. Testicular ACE (tACE), the male germinal specific isozyme, plays a crucial role in male fertilization. The amino-terminal region of this isozyme is different from that of somatic isozyme (sACE) and contains potential O-linked glycosylation sites. By multiple mutagenesis after an in silico prediction, amino acid residues acquiring O-glycans were assigned. Both GPIase and dipeptidase activities were compared between O-glycan null mutant and wild-type molecules, but no differences were found. Furthermore, the wild-type tACE was produced in two different cells (COS7 and CHO) and its activities compared. The GPIase activity, but not dipeptidase, was apparently higher for CHO-derived molecule than COS7. Sensitivity to neuraminidase and O-glycosidase digestions and the profile of glycosylation were quite different between these two molecules. Moreover, serial digestions with neuraminidase and O-glycosidase have no influence on GPIase activity of both molecules, suggesting that the sialylation and the presence of O-glycan has no influence on tACE enzyme activities, while the set of glycans modulate GPIase activity.
Kondoh
Gen
G
Laboratory of Animal Experiments for Regeneration, Institute for Frontier Medical Sciences, Kyoto University, Kyoto, Japan. kondohg@frontier.kyoto-u.ac.jp
Watanabe
Hitomi
H
Tashima
Yuko
Y
Maeda
Yusuke
Y
Kinoshita
Taroh
T
D000818:Animals; D016466:CHO Cells; D019556:COS Cells; D002522:Chlorocebus aethiops; D006224:Cricetinae; D003412:Cricetulus; D004150:Dipeptidases / Q000378:metabolism*; D017261:Glycosylphosphatidylinositols / Q000378:metabolism*; D008297:Male; D051379:Mice; D008957:Models, Genetic; D009154:Mutation; D007703:Peptidyl-Dipeptidase A / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins; D013737:Testis / Q000201:enzymology* / Q000378:metabolism; D014162:Transfection
D017261:Glycosylphosphatidylinositols; D011994:Recombinant Proteins; D004150:Dipeptidases; C038937:dipeptidase; D007703:Peptidyl-Dipeptidase A