Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, 1800 Lihu Road, Wuxi, 214122, Jiangsu, China.
School of Biotechnology, Jiangnan University, 1800 Lihu Road, Wuxi, 214122, Jiangsu, China.
Microb Cell Fact. 2019 Aug 13;18(1):136. doi: 10.1186/s12934-019-1188-8.
Microbial cell factories are widely used in the production of acidic products such as organic acids and amino acids. However, the metabolic activity of microbial cells and their production efficiency are severely inhibited with the accumulation of intracellular acidic metabolites. Therefore, it remains a key issue to enhance the acid tolerance of microbial cells. In this study, we investigated the effects of four ATP-binding cassette (ABC) transporters on acid stress tolerance in Lactococcus lactis.
Overexpressing the rbsA, rbsB, msmK, and dppA genes exhibited 5.8-, 12.2-, 213.7-, and 5.2-fold higher survival rates than the control strain, respectively, after acid shock for 3 h at pH 4.0. Subsequently, transcriptional profile alterations in recombinant strains were analyzed during acid stress. The differentially expressed genes associated with cold-shock proteins (csp), fatty acid biosynthesis (fabH), and coenzyme A biosynthesis (coaD) were up-regulated in the four recombinant strains during acid stress. Additionally, some genes were differentially expressed in specific recombinant strains. For example, in L. lactis (RbsB), genes involved in the pyrimidine biosynthetic pathway (pyrCBDEK) and glycine or betaine transport process (busAA and busAB) were up-regulated during acid stress, and the argG genes showed up-regulations in L. lactis (MsmK). Finally, we found that overexpression of the ABC transporters RbsB and MsmK increased intracellular ATP concentrations to protect cells against acidic damage in the initial stage of acid stress. Furthermore, L. lactis (MsmK) consistently maintained elevated ATP concentrations under acid stress.
This study elucidates the common and specific mechanisms underlying improved acid tolerance by manipulating ABC transporters and provides a further understanding of the role of ABC transporters in acid-stress tolerance.
微生物细胞工厂广泛用于生产有机酸和氨基酸等酸性产物。然而，随着细胞内酸性代谢物的积累，微生物细胞的代谢活性及其生产效率受到严重抑制。因此，提高微生物细胞的耐酸性仍然是一个关键问题。在本研究中，我们研究了四种 ATP 结合盒（ABC）转运蛋白对乳球菌耐酸应激的影响。
在 pH 值为 4.0 的酸性冲击 3 小时后，过表达 rbsA、rbsB、msmK 和 dppA 基因的菌株的存活率分别比对照菌株高 5.8、12.2、213.7 和 5.2 倍。随后，分析了重组菌株在酸胁迫下的转录谱变化。在四种重组菌株中，与冷休克蛋白（csp）、脂肪酸生物合成（fabH）和辅酶 A 生物合成（coaD）相关的差异表达基因在酸胁迫下上调。此外，一些基因在特定的重组菌株中差异表达。例如，在乳球菌（RbsB）中，嘧啶生物合成途径（pyrCBDEK）和甘氨酸或甜菜碱转运过程（busAA 和 busAB）相关基因在酸胁迫下上调，argG 基因在乳球菌（MsmK）中上调。最后，我们发现 ABC 转运蛋白 RbsB 和 MsmK 的过表达增加了细胞内 ATP 浓度，从而在酸性应激的初始阶段保护细胞免受酸性损伤。此外，乳球菌（MsmK）在酸性胁迫下始终保持较高的 ATP 浓度。
本研究阐明了操纵 ABC 转运蛋白提高耐酸性的共同和特定机制，进一步了解了 ABC 转运蛋白在耐酸应激中的作用。
