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臭味假单胞菌乙酰乙酰辅酶A硫解酶的提纯及性质研究
引用本文:李钦,L.Nicholas Ornston.臭味假单胞菌乙酰乙酰辅酶A硫解酶的提纯及性质研究[J].中国生物化学与分子生物学报,1989,5(6):509-514.
作者姓名:李钦  L.Nicholas Ornston
作者单位:中国科学院微生物研究所 北京 (李钦),耶鲁大学生物系 美国纽黑文(L.Nicholas Ornston)
摘    要:从臭味假单胞菌中提纯97倍的AcAcCoA硫解酶在聚丙烯酰胺凝胶电泳上是均一的一带。该酶分子量为170,000,每分子含有4个亚基,亚基分子量为42,000。该酶的等电点为pI6.7。它的N-末端为丙氨酸,N-末端是单一的。该酶催化反应的Km值为10.2μmol/L,最大反应速度为16.7μmol/min·mg。 臭味假单胞菌细胞粗提液透析后,经DEAE-纤维素(DE-52)柱色谱,从洗脱液中可同时得到四个酶的活力峰:乙酰乙酸琥珀酰辅酶A转移酶,AcAcCoA硫解酶,β-酮已二酸琥珀酰辅酶A转移酶和β-酮己二酸单酰辅酶A硫解酶。一般认为在细菌的芳径代谢中存在β-酮己二酸代谢途径,上述四个酶的活力峰同时存在说明除β-酮已二酸代谢途径外,还同时存在乙酰乙酸代谢途径。

关 键 词:臭味假单胞菌  乙酰乙酰辅酶A  硫解酶  
收稿时间:1989-12-20

Purification and Properties of Acetoacetyl-Coenzyme A Thiolase From Pseudomonas Putida
Li,Qin.Purification and Properties of Acetoacetyl-Coenzyme A Thiolase From Pseudomonas Putida[J].Chinese Journal of Biochemistry and Molecular Biology,1989,5(6):509-514.
Authors:Li  Qin
Institution:(Institute of Microbiology, Academia Sinica, Beijing) L. Nicholas Ornston (Department of Biology, Yale University, New Haven, CT 06520, USA
Abstract:The acetoacetyl-coenzyme A thiolase (AcAcCoA thiolase) of Peudomo-nas putida has been purified 97-fold to homogeneity as judged by polyacrylamide gel electrophoresis at pH8.3 with different concentration gels. The enzyme has a molecular weight of 170,000, and is composed of four identical subunits. The subunit molecular weight is 42,000. The isoelectric point of the AcAcCoA thiolase obtained by gel electrofocusing was pI6.7. Alanine was the sole N-terminal amino acid detected by dansylation. The thiolase catalyzes the CoA dependent cleavage of AcAcCoA into two molecules of AcCoA. In the direction of AcAcCoA cleavage, the maximal velocity (Vmax) was 16.7μmol/min.mg, and Km value was 10.2μmol/L.The dialyzed crude extract of Pseudomonas putida cells upon DEAE-cellulose (DE-52) column chromatography gave rise to four peaks with activities representing four different enzymes: acetoacetate succinyl-CoA transferase, AcAcCoA thiolase, β-ketoadipate succinyl-CoA transferase, and β-ketoadipyl-CoA thiolase. It appeared that the β-ketoadipate pathway and the acetoacetate pathway existed simultaneously in Pseudomonas putida.
Keywords:Pseudomonas' putida Acetoacetyl coenzyme Thiolase
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