Improvement of Multiple-Stress Tolerance and Lactic Acid Production in Lactococcus lactis NZ9000 under Conditions of Thermal Stress by Heterologous Expression of Escherichia coli dnaK
Improvement of Multiple-Stress Tolerance and Lactic Acid Production in Lactococcus lactis NZ9000 under Conditions of Thermal Stress by Heterologous Expression of Escherichia coli dnaK
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DOI：
10.1128/aem.02878-09
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发表时间：
2010-07-01
影响因子：
4.4
通讯作者：
Sonomoto, Kenji
中科院分区：
文献类型：
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作者：
Abdullah-Al-Mahin;Sugimoto, Shinya;Sonomoto, Kenji
研究了乳酸链球菌素诱导的dNAK在乳酸乳球菌中的表达，结果表明该表达可以提高乳酸发酵的效率和抗逆性。利用乳链菌素诱导表达系统，在乳酸乳杆菌NZ9000中表达了乳酸乳杆菌(DNAK(11a))和大肠埃希氏菌(DNAK(Eco))的DNAK蛋白。与含有空载体pNZ8048(命名为NZ-VECTOR)和表达dNAK(11a)的菌株(命名为NZ-LDNAK)相比，表达dNAK(Eco)的菌株(命名为NZ-EDNAK)在GM17液体介质中对40℃的热胁迫表现出更强的耐受性。热处理6h后，细胞存活率下降4.6倍。然而，在这些条件下，NZ-EDNAK的活力提高了13.5倍，DNAK(Eco)的产生浓度非常低。虽然异源表达的DNAK(Eco)对DNAK(11a)的产量没有影响，但热处理使NZ-VECTOR和NZ-EDNAK的DNAK(11a)水平分别增加了3.5和3.6倍。此外，NZ-EDNAK还表现出对包括3%氯化钠、5%乙醇和0.5%乳酸(pH 5.47)在内的多重胁迫的耐受性。在CMG培养基中，在30℃时，NZ-EDNAK的乳酸产量和最大乳酸产量高于NZ-Vector30℃时的相应值。有趣的是，在40℃时，NZ-EDNAK的这些值与对照菌株在30℃时的相应值没有显著差异。这些发现表明，dNAK(Eco)的异源表达增强了蛋白质和酶的质量控制，从而改善了高温下的生长和乳酸发酵。
The effects of nisin-induced dnaK expression in Lactococcus lactis were examined, and this expression was shown to improve stress tolerance and lactic acid fermentation efficiency. Using a nisin-inducible expression system, DnaK proteins from L. lactis (DnaK(Lla)) and Escherichia coli (DnaK(Eco)) were produced in L. lactis NZ9000. In comparison to a strain harboring the empty vector pNZ8048 (designated NZ-Vector) and one expressing dnaK(Lla) (designated NZ-LDnaK), the dnaK(Eco)-expressing strain, named NZ-EDnaK, exhibited more tolerance to heat stress at 40 degrees C in GM17 liquid medium. The cell viability of NZ-Vector was reduced 4.6-fold after 6 h of heat treatment. However, NZ-EDnaK showed 13.5-fold increased viability under these conditions, with a very low concentration of DnaK(Eco) production. Although the heterologous expression of dnaK(Eco) did not effect DnaK(Lla) production, heat treatment increased the DnaK(Lla) level 3.5- and 3.6-fold in NZ-Vector and NZ-EDnaK, respectively. Moreover, NZ-EDnaK showed tolerance to multiple stresses, including 3% NaCl, 5% ethanol, and 0.5% lactic acid (pH 5.47). In CMG medium, the lactate yield and the maximum lactate productivity of NZ-EDnaK were higher than the corresponding values for NZ-Vector at 30 degrees C. Interestingly, at 40 degrees C, these values of NZ-EDnaK were not significantly different from the corresponding values for the control strain at 30 degrees C. Lactate dehydrogenase (LDH) activity was also found to be stable at 40 degrees C in the presence of DnaK(Eco). These findings suggest that the heterologous expression of dnaK(Eco) enhances the quality control of proteins and enzymes, resulting in improved growth and lactic acid fermentation at high temperature.