Maragoudakis M E, Strassman M
J Bacteriol. 1967 Sep;94(3):512-6. doi: 10.1128/jb.94.3.512-516.1967.
Cell-free extracts of Acetobacter suboxydans were prepared which were capable of condensing alpha-ketoisovalerate with (14)C-labeled acetyl-coenzyme A to yield (14)C-labeled alpha-isopropylmalate. The product of the reaction was isolated by paper and column chromatography and was characterized by recrystallization with synthetic alpha-isopropylmalic acid to constant specific radioactivity. The formation of alpha-isopropylmalate by extracts of A. suboxydans plus the ability of the organism to grow in a simple glucose-glycerol medium containing glutamic acid as the only amino acid indicate that the pathway for leucine biosynthesis shown to exist in yeast and Salmonella typhimurium also occurs in A. suboxydans. As a comparison, the condensation of oxalacetate and ((14)C) acetyl-coenzyme A to yield ((14)C) citric acid was shown, by similar means, to occur in A. suboxydans. This is of interest since the existence of this classical condensing enzyme has hitherto not been demonstrated in this organism. This reaction was further demonstrated in cell-free extracts of A. suboxydans by means of a spectrophotometric assay at 232 mmu which measured the cleavage of the carbon-sulfur bond of acetyl-coenzyme A in the presence of oxalacetate. Comparison of the specific activities of crude cell-free extracts indicated a much more extensive occurrence of this reaction in yeast than in A. suboxydans.
制备了弱氧化醋杆菌的无细胞提取物，该提取物能够将α-酮异戊酸与（14）C标记的乙酰辅酶A缩合，生成（14）C标记的α-异丙基苹果酸。反应产物通过纸色谱和柱色谱进行分离，并用合成的α-异丙基苹果酸重结晶至恒定的比放射性进行表征。弱氧化醋杆菌提取物形成α-异丙基苹果酸，以及该生物体在以谷氨酸作为唯一氨基酸的简单葡萄糖-甘油培养基中生长的能力，表明在酵母和鼠伤寒沙门氏菌中存在的亮氨酸生物合成途径在弱氧化醋杆菌中也存在。作为比较，通过类似的方法表明，草酰乙酸和（（14）C）乙酰辅酶A缩合生成（（14）C）柠檬酸的反应在弱氧化醋杆菌中发生。这很有趣，因为迄今为止在该生物体中尚未证明这种经典缩合酶的存在。通过在232毫微米处的分光光度测定法，在草酰乙酸存在下测量乙酰辅酶A的碳-硫键的裂解，进一步在弱氧化醋杆菌的无细胞提取物中证明了该反应。粗无细胞提取物的比活性比较表明，该反应在酵母中的发生比在弱氧化醋杆菌中更为广泛。
