Studies on pig liver mevalonate-5-diphosphate decarboxylase.
A procedure in which three sequential enzymes of cholesterol biosynthesis, mevalonate kinase (ATP: (R)-mevalonate 5-phosphotransferase, EC 2.7.1.36), phosphomevalonate kinase (ATP: (R)-5-phosphomevalonate phosphotransferase, EC 2.7.4.2) and mevalonate-5-diphosphate decarboxylase (ATP: (R)-5-diphosphomevalonate carboxy-lyase (dehydrating), EC 4.1.1.33), from pig liver, could be purified in the one operation is described. Mevalonate kinase and phosphomevalonate kinase were utilized for the enzymic synthesis of mevalonate 5-diphosphate (both 1-14C-labelled and unlabelled), the substrate for mevalonate-5-diphosphate decarboxylase, using excess free ATP4-. A radioactive assay for the enzyme, based on the release of 14CO2 from [1-14C]mevalonate-5-diphosphate, was developed. The assay allowed reassessment of the metal and nucleotide specificity of the decarboxylase. ATP could be partially replaced by GTP and ITP, but no activity was observed with CTP, UTP or TTP. Apparent activation of the enzyme by ATP4- was observed as found for mevalonate kinase (C.S. Lee and W.J. O'Sullivan (1983) Biochim. Biophys. Acta 747, 215-224) and phosphomevalonate kinase (C.S. Lee and W.J. O'Sullivan (1985) Biochim. Biophys. Acta 839, 83-89). The presence of 1 mM excess free ATP4-, above that complexed as the substrate MgATP2-, decreased the Km for MgATP2- from 0.45 mM to 0.15 mM. MgADP- was shown to act as a competitive inhibitor with respect to MgATP2-.
Chiew
Y E
YE
School of Biochemistry, University of New South Wales, Kensington, Australia.
O'Sullivan
W J
WJ
Lee
C S
CS
D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism; D000818:Animals; D002262:Carboxy-Lyases / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D008670:Metals / Q000378:metabolism; D008798:Mevalonic Acid / Q000031:analogs & derivatives / Q000096:biosynthesis; D010770:Phosphotransferases / Q000378:metabolism; D017853:Phosphotransferases (Alcohol Group Acceptor); D017856:Phosphotransferases (Phosphate Group Acceptor); D013379:Substrate Specificity; D013552:Swine
D008670:Metals; C046285:5-diphosphomevalonic acid; D000255:Adenosine Triphosphate; D010770:Phosphotransferases; D017853:Phosphotransferases (Alcohol Group Acceptor); C021796:mevalonate kinase; D017856:Phosphotransferases (Phosphate Group Acceptor); C022371:phosphomevalonate kinase; D002262:Carboxy-Lyases; C022503:pyrophosphomevalonate decarboxylase; D008798:Mevalonic Acid