Lactobacillus buchneri and Oenococcus oeni are two unique ethanol-tolerant Gram-positive bacteria species. Genome comparison analyses revealed that L. buchneri and O. oeni possess a pntAB locus that was absent in almost all other lactic acid bacteria (LAB) genomes. Our hypothesis is that the pntAB locus contributes to the ethanol tolerance trait of these two distinct ethanol-tolerant organisms. The pntAB locus, consisting of the pntA and pntB genes, codes for NADP(H) transhydrogenase subunits. This membrane-bound transhydrogenase catalyzes the reduction of NADP+ and is known as an important enzyme in maintaining cellular redox balance. In this study, the transhydrogenase operon from L. buchneri NRRL B-30929 and O. oeni PSU-1 were cloned and analyzed. The LbpntB shared 71.0% identity with the O. oeni (OopntB). The entire pntAB locus was expressed in Lactococcus lactis ssp. lactis IL1403 resulting in an increased tolerance to ethanol (6%), butanol (1.8%) and isopropanol (1.8%) when compared to the control strain. However, the recombinant E. coli cells carrying the entire pntAB locus did not show any improved ethanol tolerance. Independent expression of OopntB and LbpntB in recombinant E. coli BL21(DE3)pLysS host demonstrated higher tolerance to ethanol when compared with a control E. coli BL21(DE3)pLysS strain carrying pET28b vector. Ethanol tolerance comparison of E. coli strains carrying LbpntB and OopntB showed that LbpntB conferred higher ethanol tolerance (4.5%) and resulted in greater biomass, while the OopntB conferred lower ethanol tolerance (4.0%) resulted lower biomass. Therefore, the pntB gene from L. buchneri is a better choice in generating higher ethanol tolerance. This is the first study to uncover the role of pntAB locus on ethanol tolerance.

中文翻译：

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与oenococcus oeni和布氏乳杆菌的pntAB基因座相关的乙醇耐受性增加。

布氏乳杆菌和oenococcus oeni是两个独特的乙醇耐受革兰氏阳性细菌。基因组比较分析显示，布氏乳杆菌和O. oeni拥有一个pntAB基因座，几乎所有其他乳酸菌（LAB）基因组中都没有。我们的假设是pntAB基因座有助于这两种不同的耐乙醇生物的耐乙醇性状。由pntA和pntB基因组成的pntAB基因座编码NADP（H）转氢酶亚基。这种膜结合的转氢酶催化NADP +的还原，是维持细胞氧化还原平衡的重要酶。在本研究中，克隆并分析了来自布氏乳杆菌NRRL B-30929和O. oeni PSU-1的转氢酶操纵子。LbpntB与O. oeni（OopntB）具有71.0％的同一性。整个pntAB基因座在乳酸乳球菌中表达。与对照菌株相比，乳酸菌IL1403可提高对乙醇（6％），丁醇（1.8％）和异丙醇（1.8％）的耐受性。但是，携带整个pntAB基因座的重组大肠杆菌细胞未显示出任何改善的乙醇耐受性。与携带pET28b载体的对照大肠杆菌BL21（DE3）pLysS菌株相比，重组大肠杆菌BL21（DE3）pLysS宿主中OopntB和LbpntB的独立表达表现出更高的乙醇耐受性。携带LbpntB和OopntB的大肠杆菌菌株的乙醇耐受性比较显示，LbpntB赋予较高的乙醇耐受性（4.5％）并导致更大的生物量，而OopntB赋予较低的乙醇耐受性（4.0％）导致较低的生物量。因此，来自L的pntB基因。布氏杆菌是产生更高乙醇耐受性的更好选择。这是第一个揭示pntAB基因座对乙醇耐受性作用的研究。