title: Characterization of a Zn(2+)-requiring glycerophosphocholine cholinephosphodiesterase possessing p-nitrophenylphosphocholine phosphodiesterase activity.
abstract: p-Nitrophenylphosphocholine phosphodiesterase activity was purified 5000-fold from mouse brain by treatment of membranes with Bacillus cereus phospholipase C preparation and sequential chromatographies on concanavalin A-Sepharose and CM-Sephadex columns. The phosphodiesterase (Zn(2+)-requiring) showed Km and Vmax. values of 5.5 microM and 4.2 mumol/min per mg respectively in the hydrolysis of p-nitrophenylphosphocholine, and possessed an optimum pH of 10.5 and a molecular mass of approx. 74 kDa. The purified enzyme was found to convert glycerophosphocholine into glycerol and phosphocholine, with Km and Vmax. of 48 microM and 5 mumol/min per mg respectively. In the hydrolysis of glycerophosphocholine the enzyme also exhibited a Zn2+ requirement and optimal pH at 10.5. Additionally, the p-nitrophenylphosphocholine phosphodiesterase activity was competitively inhibited by glycerophosphocholine, with a Ki value of 50 microM. These observations, together with chromatographic behaviour and heat-denaturation analyses, indicate that both p-nitrophenylphosphocholine phosphodiesterase and glycerophosphocholine cholinephosphodiesterase activities reside in the same protein.
authors.affiliation: Department of Food and Nutrition, Chungnam National University, Korea.
authors.forename: D E
authors.initials: DE
authors.lastname: Sok
authors.forename: M R
authors.initials: MR
authors.lastname: Kim
mesh_terms: D000818:Animals; D001921:Brain / Q000201:enzymology; D002413:Cations, Divalent; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D006358:Hot Temperature; D007700:Kinetics; D051379:Mice; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism*; D015032:Zinc / Q000378:metabolism*
chemical_list: D002413:Cations, Divalent; D010727:Phosphoric Diester Hydrolases; C061931:glycerophosphocholine cholinephosphodiesterase; D015032:Zinc