Technische Universität München, Lehrstuhl Technische Mikrobiologie, Freising, Germany.
Food Microbiol. 2011 Feb;28(1):29-37. doi: 10.1016/j.fm.2010.08.001. Epub 2010 Aug 14.
Lactobacillus sanfranciscensis is the key bacterium in traditional sourdough fermentation. The molecular background of its oxygen tolerance was investigated by comparison of wild type and NADH-oxidase (Nox) knock out mutants. The nox gene of L. sanfranciscensis DSM20451(T) coding for a NADH-oxidase (Nox) was inactivated by single crossover integration to yield strain L. sanfranciscensis DSM20451Δnox. By inactivation of the native NADH-oxidase gene, it was ensured that besides fructose, O(2) can react as an electron acceptor. In aerated cultures the mutant strain was only able to grow in MRS media supplemented with fructose as electron acceptor, whereas the wild type strain showed a fructose independent growth response. The use of oxygen as an external electron acceptor enables L. sanfranciscensis to shift from acetyl-phosphate into the acetate branch and gain an additionally ATP, while the reduced cofactors were regenerated by Nox-activity. In aerated cultures the wild type strain formed a fermentation ratio of lactate to acetate of 1.09 in MRS supplemented with fructose after 24 h of fermentation, while the mutant strain formed a fermentation ratio of 3.05. Additionally, L. sanfranciscensis showed manganese-dependent growth response in aerated cultures, the final OD and growth velocity was increased in media supplemented with manganese. The expression of two predicted Mn(2+)/Fe(2+) transporters MntH1 and MntH2 in L. sanfranciscensis DSM20451(T) was verified by amplification of a 318 bp fragment of MntH1 and a 239 bp fragment of MntH2 from cDNA library obtained from aerobically, exponentially growing cells of L. sanfranciscensis DSM20451(T) in MRS. Moreover, the mutant strain DSM20451Δnox was more sensitive to the superoxide generating agent paraquat and showed inhibition of growth on diamide-treated MRS-plates without fructose supplementation.
旧金山乳杆菌是传统酸面团发酵中的关键细菌。通过比较野生型和 NADH 氧化酶（Nox）敲除突变体，研究了其耐氧性的分子背景。通过单交换整合失活了编码 NADH 氧化酶（Nox）的 L. sanfranciscensis DSM20451（T）的 nox 基因，得到了 L. sanfranciscensis DSM20451Δnox 突变株。通过失活天然 NADH 氧化酶基因，确保除了果糖外，O（2）可以作为电子受体进行反应。在通气培养中，突变株只能在补充果糖作为电子受体的 MRS 培养基中生长，而野生型菌株则表现出果糖非依赖性生长反应。利用氧气作为外部电子受体，使 L. sanfranciscensis 能够从乙酰磷酸转移到乙酸分支，并获得额外的 ATP，同时通过 Nox 活性再生还原辅酶。在通气培养中，野生型菌株在补充果糖的 MRS 培养基中发酵 24 小时后形成的乳酸与乙酸的发酵比为 1.09，而突变株形成的发酵比为 3.05。此外，L. sanfranciscensis 在通气培养中表现出锰依赖性生长反应，在补充锰的培养基中最终 OD 和生长速度增加。通过从 L. sanfranciscensis DSM20451（T）的好氧、指数生长期的 cDNA 文库中扩增出 318 bp 的 MntH1 片段和 239 bp 的 MntH2 片段，验证了 L. sanfranciscensis DSM20451（T）中两个预测的 Mn（2+）/Fe（2+）转运蛋白 MntH1 和 MntH2 的表达。此外，突变株 DSM20451Δnox 对超氧化物生成剂百草枯更为敏感，并表现出在没有果糖补充的情况下，在添加二酰胺的 MRS 平板上生长受到抑制。
