STUDIES ON PIG-LIVER MEVALONATE-5-DIPHOSPHATE DECARBOXYLASE
STUDIES ON PIG-LIVER MEVALONATE-5-DIPHOSPHATE DECARBOXYLASE
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DOI：
10.1016/0167-4838(87)90170-1
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发表时间：
1987-12-18
期刊：
影响因子：
--
通讯作者：
LEE, CS
中科院分区：
文献类型：
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作者：
CHIEW, YE;OSULLIVAN, WJ;LEE, CS
描述了一种从猪肝中纯化胆固醇生物合成的三种连续酶甲羟戊酸激酶（ATP：（R）-甲羟戊酸5-磷酸转移酶，EC www.example.com）、磷酸甲羟戊酸激酶（ATP：（R）-5-磷酸甲羟戊酸磷酸转移酶，EC www.example.com）和甲羟戊酸-5-二磷酸脱羧酶（ATP：（R）-5-二磷酸甲羟戊酸羧基裂解酶（脱水），EC www.example.com）的方法。甲羟戊酸激酶和磷酸甲羟戊酸激酶用于使用过量的游离ATP 4-酶促合成甲羟戊酸-5-二磷酸（1 - 14C-标记和未标记），甲羟戊酸-5-二磷酸脱羧酶的底物。基于从[1 - 14C]甲羟戊酸-5-二磷酸释放14CO2，开发了酶的放射性测定。该测定允许重新评估脱羧酶的金属和核苷酸特异性。ATP可被GTP和ITP部分替代，而CTP、UTP和TTP则无活性。与甲羟戊酸激酶（C. S. Lee和W.J. O''Sullivan（1983）Biochim. Biophys. Acta 747，215 - 224）和磷酸甲羟戊酸激酶（C.S. Lee和W.J. O''Sullivan（1985）Biochim. Biophys. Acta 839，83 - 89）。过量的1 mM游离ATP 4-的存在下，以上的复合作为底物MgATP 2-，降低了米氏为MgATP 2-从0.45 mM至0.15 mM。MgADP-被证明是作为一个竞争性抑制剂相对于MgATP 2-。
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-.