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Enhanced 23-Butanediol Production by Optimizing Fermentation Conditions and Engineering Klebsiella oxytoca M1 through Overexpression of Acetoin Reductase

机译：通过优化发酵条件和通过乙酰丙酮还原酶的过表达工程化产克雷伯氏菌M1来增强23-丁二醇的生产

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摘要

Microbial production of 2,3-butanediol (2,3-BDO) has been attracting increasing interest because of its high value and various industrial applications. In this study, high production of 2,3-BDO using a previously isolated bacterium Klebsiella oxytoca M1 was carried out by optimizing fermentation conditions and overexpressing acetoin reductase (AR). Supplying complex nitrogen sources and using NaOH as a neutralizing agent were found to enhance specific production and yield of 2,3-BDO. In fed-batch fermentations, 2,3-BDO production increased with the agitation speed (109.6 g/L at 300 rpm vs. 118.5 g/L at 400 rpm) along with significantly reduced formation of by-product, but the yield at 400 rpm was lower than that at 300 rpm (0.40 g/g vs. 0.34 g/g) due to acetoin accumulation at 400 rpm. Because AR catalyzing both acetoin reduction and 2,3-BDO oxidation in K. oxytoca M1 revealed more than 8-fold higher reduction activity than oxidation activity, the engineered K. oxytoca M1 overexpressing the budC encoding AR was used in fed-batch fermentation. Finally, acetoin accumulation was significantly reduced by 43% and enhancement of 2,3-BDO concentration (142.5 g/L), yield (0.42 g/g) and productivity (1.47 g/L/h) was achieved compared to performance with the parent strain. This is by far the highest titer of 2,3-BDO achieved by K. oxytoca strains. This notable result could be obtained by finding favorable fermentation conditions for 2,3-BDO production as well as by utilizing the distinct characteristic of AR in K. oxytoca M1 revealing the nature of reductase.
机译：由于其高价值和各种工业应用，微生物生产2,3-丁二醇（2,3-BDO）引起了越来越多的兴趣。在这项研究中，通过优化发酵条件和过表达乙酰辅酶还原酶（AR），利用先前分离的细菌产酸克雷伯菌（Klebsiella oxytoca M1）高产了2,3-BDO。发现提供复杂的氮源并使用NaOH作为中和剂可以提高2,3-BDO的比产量和收率。在分批补料发酵中，2,3-BDO的产生随着搅拌速度的增加而增加（300 rpm时为109.6 g / L，而400 rpm时为118.5 g / L），并且副产物的形成显着减少，但在400时的产量rpm低于300 rpm（0.40 g / g与0.34 g / g），这是因为乙酰肽在400 rpm时积累。因为AR催化催产假单胞菌M1中的丙酮酸还原和2,3-BDO氧化显示还原活性比氧化活性高8倍以上，所以过表达编码AR的budC的工程化的催产假单胞菌M1用于分批补料发酵。最终，与苯磺酸盐相比，丙酮的累积量显着降低了43％，并提高了2,3-BDO浓度（142.5 g / L），产量（0.42 g / g）和生产率（1.47 g / L / h）。亲本菌株。这是迄今为止产氧克雷伯氏菌菌株达到的最高的2,3-BDO滴度。通过发现有利的发酵条件以生产2,3-BDO以及利用催产假单胞菌M1中AR的独特特性（揭示还原酶的性质），可以获得明显的结果。

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