STUDIES ON CELL-FREE METABOLISM - ETHANOL-PRODUCTION BY A YEAST GLYCOLYTIC SYSTEM RECONSTITUTED FROM PURIFIED ENZYMES
STUDIES ON CELL-FREE METABOLISM - ETHANOL-PRODUCTION BY A YEAST GLYCOLYTIC SYSTEM RECONSTITUTED FROM PURIFIED ENZYMES
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DOI：
10.1016/0168-1656(85)90029-x
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发表时间：
1985-01-01
影响因子：
4.1
通讯作者：
SCOPES, RK
中科院分区：
文献类型：
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作者：
WELCH, P;SCOPES, RK
从单独纯化的酶建立了重建的糖酵解系统，以模拟酵母[酿酒酵母]将葡萄糖转化为乙醇和CO2。如果葡萄糖的输入与ATP降解速率适当匹配，则发生持续和广泛的转化。ATP酶的活性可以被砷酸盐所取代，砷酸盐将ATP合成与糖酵解分开。解偶联的模式进行了研究，并得出结论，人工中间体，1-砷-3-磷酸甘油酸，具有不超过几毫秒的半衰期。4 mM浓度的砷酸盐可以模拟相当于10 μ mol ml-1 min-1的ATP酶活性。重组酶系统能够在8 h内完全降解1 M（18%wt/vol）葡萄糖，得到9%（wt/vol）乙醇。测量代谢过程中代谢物的水平，以检测限速步骤。重组酶系统的成功操作表明，它是可能的，在体外进行复杂的化学转化与多种酶系统。
A reconstituted glycolytic system was established from individually purified enzymes to simulate the conversion of glucose to ethanol plus CO2 by yeast [Saccharomyces cerevisiae]. Sustained and extensive conversion occurred provided thant input of glucose matched the rate of ATP degradation appropriately. ATPase activity could be replaced by arsenate, which uncoupled ATP synthesis from glycolysis. The mode of uncoupling was investigated, and it was concluded that the artificial intermediate, 1-arseno-3-phosphoglycerate, has a half-life of no more than a few milliseconds. Arsenate at 4 mM concentration could simulate the equivalent of 10 .mu.mol ml-1 min-1 of ATPase activity. The reconstituted enzyme system was capable of totally degrading 1 M (18% wt/vol) glucose in 8 h giving 9% (wt/vol) ethanol. The levels of metabolites during metabolism were measured to detect rate-limiting steps. The successful operation of the reconstituted enzyme system demonstrates that it is possible to carry out complex chemical transformations with multiple enzyme systems in vitro.