Submitochondrial localization and partial purification of the succinylCoA: 3-hydroxy-3-methylglutarate coenzyme A transferase from rat liver

Renzo Deana, Fernanda Rigoni, Arianna Donella Deana, Lauro Galzigna
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引用次数: 3

Abstract

The presence and the localization of the enzyme catalyzing the transfer of a coenzyme A molecule from succinylCoA to 3-hydroxy-3-methylglutarate has been established in rat liver mitochondria. The enzyme was found mainly in the mitochondrial matrix but some activity was also found in the inner membrane fraction. The enzyme has been purified about 100-fold from sonically-disrupted mitochondria by high-speed centrifugation, DEAE-cellulose chromatography, (NH4)2SO4 precipitation and Sephadex G-100 filtration. The enzymatic activity was recovered in the final step as a single peak. The coenzyme A transferase appears to have a molecular increases the activity and improves its stability. The enzyme is different from the succinylCoA: 3-oxoacids coenzyme A transferase and is active also on malonylCoA. The apparent Km values obtained for succinylCoA, malnyCoA and 3-hydroxy-3-methylglutarate were 2.2 · 10−4 M, 3.7 · 10−4 M and 1.7 · 10−3 M, respectively. Acetoacetate, which is the final product of the mitochondrial metabolism of hydroxy-methylglutarylCoA, showed an inhibitory effect on the enzyme activity with a Ki of 0.5 mM. The physiological role of the enzyme is discussed.

大鼠肝脏琥珀酰辅酶A: 3-羟基-3-甲基戊二酸辅酶A转移酶的亚线粒体定位和部分纯化
催化辅酶a分子从琥珀酰辅酶a向3-羟基-3-甲基戊二酸转移的酶在大鼠肝脏线粒体中的存在和定位已被证实。该酶主要存在于线粒体基质中,但在内膜部分也有活性。该酶通过高速离心、deae -纤维素层析、(NH4)2SO4沉淀和Sephadex G-100过滤从超声破坏的线粒体中纯化约100倍。酶活性在最后一步恢复为单峰。辅酶A转移酶似乎有一个分子增加活性和改善其稳定性。该酶与琥珀酰辅酶A: 3-氧酸辅酶A转移酶不同,对丙二酸辅酶A也有活性。琥珀酰辅酶a、麦芽糖辅酶a和3-羟基-3-甲基戊二酸盐的表观Km值分别为2.2·10−4 M、3.7·10−4 M和1.7·10−3 M。乙酰乙酸是羟甲基戊二酰辅酶a线粒体代谢的最终产物,在Ki为0.5 mM时对酶活性有抑制作用,并讨论了酶的生理作用。
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