Continuous production of (R)-phenylacetylcarbinol in an enzyme-membrane reactor using a potent mutant of pyruvate decarboxylase from Zymomonas mobilis.
Continuous production of (R)-phenylacetylcarbinol in an enzyme-membrane reactor using a potent mutant of pyruvate decarboxylase from Zymomonas mobilis.
复制标题
使用来自运动发酵单胞菌的丙酮酸脱羧酶的有效突变体在酶膜反应器中连续生产 (R)-苯乙酰基甲醇。
DOI：
10.1002/bit.1122
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发表时间：
2001
影响因子：
3.8
通讯作者：
M. Pohl
中科院分区：
文献类型：
--
作者：
G. Goetz;P. Iwan;B. Hauer;M. Breuer;M. Pohl
对合成L-麻黄碱的中间体（R）-苯乙酰基甲醇（PAC）的连续酶促反应进行了优化。我们比较了三个丙酮酸脱羧酶（PDC），PDC从酿酒酵母，PDC从运动发酵单胞菌，后者的一个有效的突变体，PDCW 392 M，相对于它们的应用程序中的生物转化乙醛和苯甲醛为底物的适用性。其中，突变体酶是活性最高、最稳定的酶。通过测定两种醛的不同底物浓度的初始速率，研究了碳化反应的反应条件。从得到的数据推断动力学模型，其拟合实验数据具有足够的可靠性，以推断酶促过程的两种底物的最佳浓度。结果表明，使用连续反应系统和以等摩尔浓度进料两种醛最有效地进行碳化。使用连续操作的酶膜反应器的初步研究得到（R）-PAC，时空产率为81 g L（-1）。d（-1），使用50 mM的两种醛的底物浓度。酶-膜反应器的改进有利于提高产率。新的策略允许在水反应体系中从廉价的底物合成（R）-PAC。因此，它克服了严重限制当前发酵过程的副产物形成的限制。
The optimization of a continuous enzymatic reaction yielding (R)-phenylacetylcarbinol (PAC), an intermediate of the L-ephedrine synthesis, is presented. We compare the suitability of three pyruvate decarboxylases (PDC), PDC from Saccharomyces cerevisiae, PDC from Zymomonas mobilis, and a potent mutant of the latter, PDCW392M, with respect to their application in the biotransformation using acetaldehyde and benzaldehyde as substrates. Among these, the mutant enzyme was the most active and most stable one. The reaction conditions of the carboligation reaction were investigated by determining initial rate velocities with varying substrate concentrations of both aldehydes. From the resulting data a kinetic model was inferred which fits the experimental data with sufficient reliability to deduce the optimal concentrations of both substrates for the enzymatic process. The results demonstrate that the carboligation is most efficiently performed using a continuous reaction system and feeding both aldehydes in equimolar concentration. Initial studies using a continuously operated enzyme-membrane reactor gave (R)-PAC with a space-time yield of 81 g L(-1). d(-1) using a substrate concentration of 50 mM of both aldehydes. The yield was easily increased by cascadation of enzyme-membrane reactors. The new strategy allows the synthesis of (R)-PAC from cheap substrates in an aqueous reaction system. It thereby overcomes the limitation of by-product formation that severely limits the current fermentative process.