Phosphotransbutyrylase from Clostridium acetobutylicum ATCC 824 and its role in acidogenesis.

摘要: Phosphotransbutyrylase (phosphate butyryltransferase [EC 2.3.1.19]) from Clostridium acetobutylicum ATCC 824 was purified approximately 200-fold to homogeneity with a yield of 13%. Steps used in the purification procedure were fractional precipitation (NH4)2SO4, Phenyl Sepharose CL-4B chromatography, DEAE-Sephacel high-pressure liquid chromatography an anion-exchange column, and hydrophobic-interaction column. Gel filtration denaturing gel electrophoresis data consistent native enzyme having eight 31,000-molecular-weight subunits. Within physiological range pH 5.5 7, very sensitive change butyryl phosphate-forming direction showed virtually no activity below 6. This finding indicates that internal may be one important factor regulation enzyme. The less reverse direction. could use number substrates addition coenzyme A (butyryl-CoA) but had highest relative butyryl-CoA, isovaleryl-CoA, valeryl-CoA. Km values at 30 degrees C 8.0 for phosphate, CoASH (reduced form CoA) 0.11, 14, 0.26, 0.077 mM, respectively. Results product inhibition studies random Bi binding mechanism which phosphate binds more than site.