Overexpression of the phosphofructokinase encoding gene is crucial for achieving high production of D-lactate in Corynebacterium glutamicum under oxygen deprivation
Overexpression of the phosphofructokinase encoding gene is crucial for achieving high production of D-lactate in Corynebacterium glutamicum under oxygen deprivation
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DOI：
10.1007/s00253-015-6546-9
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发表时间：
2015-06-01
影响因子：
5
通讯作者：
Inui, Masayuki
中科院分区：
文献类型：
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作者：
Tsuge, Yota;Yamamoto, Shogo;Inui, Masayuki
我们先前报道了在缺氧条件下，编码葡萄糖激酶(GLK)、甘油醛磷酸脱氢酶(GAPDH)、磷酸果糖激酶(PFK)、磷酸丙糖异构酶(TPI)和二磷酸缩醛酶(FBA)的糖酵解途径的单个基因过表达对谷氨酸棒杆菌D-乳酸产量的影响。为了寻找协同作用，在目前的研究中，我们以分步的方式同时过表达了五个糖酵解基因，以评估累积过表达的糖酵解基因对D-乳酸合成的影响。有趣的是，最终的D-乳酸浓度明显不同，这取决于编码PFK的基因与其他糖酵解基因结合时是否过度表达。GLK、GAPDH、TPI和FBA编码基因的同时过表达导致D-乳酸的初始浓度在10h达到最高水平，但当浓度达到1300 mM时，D-乳酸的产生显著减慢，通常在32h后。相反，高表达PFK编码基因和GLK、GAPDH、TPI和FBA编码基因的菌株在10h的初始D-乳酸浓度比高表达GLK、GAPDH、TPI和FBA基因的菌株低12.7%。然而，该重组菌在32h后仍能产生D-乳酸，在矿物盐培养80h后达到2169 mm，这些结果表明，编码PFK的基因的过表达是实现D-乳酸高产的必要条件。我们的发现为探索利用谷氨酸杆菌在工业规模上低成本生产D-乳酸提供了有趣的选择。
We previously reported on the impacts of the overexpression of individual genes of the glycolytic pathway encoding glucokinase (GLK), glyceraldehyde phosphate dehydrogenase (GAPDH), phosphofructokinase (PFK), triosephosphate isomerase (TPI), and bisphosphate aldolase (FBA) on D-lactate productivity in Corynebacterium glutamicum under oxygen-deprived conditions. Searching for synergies, in the current study, we simultaneously overexpressed the five glycolytic genes in a stepwise fashion to evaluate the effect of the cumulative overexpression of glycolytic genes on D-lactate production. Interestingly, the final D-lactate concentration markedly differed depending on whether or not the PFK encoding gene was overexpressed when combined with overexpressing other glycolytic genes. The simultaneous overexpression of the GLK, GAPDH, TPI, and FBA encoding genes led to the highest initial D-lactate concentration at 10 h. However, this particular recombinant strain dramatically slowed producing D-lactate when a concentration of 1300 mM was reached, typically after 32 h. In contrast, the strain overexpressing the PFK encoding gene together with the GLK, GAPDH, TPI, and FBA encoding genes showed 12.7 % lower initial D-lactate concentration at 10 h than that observed with the strain overexpressing the genes coding for GLK, GAPDH, TPI, and FBA. However, this recombinant strain continued to produce D-lactate after 32 h, reaching 2169 mM after a mineral salts medium bioprocess incubation period of 80 h. These results suggest that overexpression of the PFK encoding gene is essential for achieving high production of D-lactate. Our findings provide interesting options to explore for using C. glutamicum for cost-efficient production of D-lactate at the industrial scale.