Agmatine deiminase from cucumber seedlings is a mono-specific enzyme: purification and characteristics.
Agmatine deiminase was purified to homogeneity from cucumber seedlings. The purification procedures included treatment with DE52, ammonium sulfate precipitation, DE52 column chromatography, Superdex 200 column chromatography, and agmatine-(CNBr)-diaminohexane-CNBr-activated-Sepharose 4B column chromatography. The purified agmatine deiminase exhibited a specific activity of 242nkat/mg protein at 30 degrees C, pH 7.0, with a yield of 33%. The molecular mass of the native enzyme was 67kDa, as estimated by Superdex 200 column chromatography. On the other hand, SDS-PAGE showed that the molecular masses of the subunits with 1% SDS and 5% of 2-mercaptoethanol treatment and with additional N-glycosidase F treatment were 47 and 36kDa, respectively. These results suggest that agmatine deiminase from cucumber is a glycoprotein. The Km of the enzyme for agmatine was 16microM and arcaine was a potent competitive inhibitor of the enzyme, with a Ki of 7.1microM. The enzyme was stable for 2 months at 4 degrees C. The enzyme does not have putrescine synthase activity or the activities of its components ornithine and putrescine transcarbamylase. The characteristics of the enzyme purified from cucumber were like those of the enzyme from maize. These results indicate that agmatine deiminase is distinctly different from putrescine synthase in higher plants.
Sakakibara
Yoko
Y
Course of Science Education, Aichi University of Education, Igaya, Kariya 448-8542, Japan.
Yanagisawa
Hiroshi
H
D001645:Biguanides / Q000494:pharmacology; D002850:Chromatography, Gel; D018553:Cucumis sativus / Q000201:enzymology*; D004591:Electrophoresis, Polyacrylamide Gel; D006867:Hydrolases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D007700:Kinetics; D008970:Molecular Weight; D009097:Multienzyme Complexes / Q000378:metabolism; D009954:Ornithine Carbamoyltransferase / Q000378:metabolism; D017851:Phosphotransferases (Carboxyl Group Acceptor) / Q000378:metabolism; D018520:Plant Shoots / Q000201:enzymology; D036226:Seedlings / Q000201:enzymology*; D013379:Substrate Specificity; D014166:Transferases / Q000378:metabolism
D001645:Biguanides; D009097:Multienzyme Complexes; C006624:arcaine; C031867:putrescine synthase; D014166:Transferases; D009954:Ornithine Carbamoyltransferase; D017851:Phosphotransferases (Carboxyl Group Acceptor); D006867:Hydrolases; C019626:agmatine deiminase