Department of Botany and Microbiology, The University of Oklahoma, Norman, OK 73019, USA.
Department of Civil and Environmental Engineering, University of Washington, Seattle, WA 98195, USA.
Microbiology (Reading). 2011 Oct;157(Pt 10):2912-2921. doi: 10.1099/mic.0.051284-0. Epub 2011 Jul 28.
Syntrophic growth involves the oxidation of organic compounds and subsequent transfer of electrons to an H(2)- or formate-consuming micro-organism. In order to identify genes involved specifically in syntrophic growth, a mutant library of Desulfovibrio alaskensis G20 was screened for loss of the ability to grow syntrophically with Methanospirillum hungatei JF-1. A collection of 20 mutants with an impaired ability to grow syntrophically was obtained. All 20 mutants grew in pure culture on lactate under sulfidogenic conditions at a rate and to a maximum OD(600) similar to those of the parental strain. The largest number of mutations that affected syntrophic growth with lactate was in genes encoding proteins involved in H(2) oxidation, electron transfer, hydrogenase post-translational modification, pyruvate degradation and signal transduction. The qrcB gene, encoding a quinone reductase complex (Qrc), and cycA, encoding the periplasmic tetrahaem cytochrome c(3) (TpIc(3)), were required by G20 to grow syntrophically with lactate. A mutant in the hydA gene, encoding an Fe-only hydrogenase (Hyd), is also impaired in syntrophic growth with lactate. The other mutants grew more slowly than the parental strain in syntrophic culture with M. hungatei JF-1. qrcB and cycA were shown previously to be required for growth of G20 pure cultures with H(2) and sulfate. Washed cells of the parental strain produced H(2) from either lactate or pyruvate, but washed cells of qrcB, cycA and hydA mutants produced H(2) at rates similar to the parental strain from pyruvate and did not produce significant amounts of H(2) from lactate. Real-time quantitative PCR assays showed increases in expression of the above three genes during syntrophic growth compared with pure-culture growth with lactate and sulfate. Our work shows that Hyd, Qrc and TpIc(3) are involved in H(2) production during syntrophic lactate metabolism by D. alaskensis G20 and emphasizes the importance of H(2) production for syntrophic lactate metabolism in this strain.
共代谢生长涉及有机化合物的氧化，随后电子被转移到消耗 H(2) 或甲酸盐的微生物。为了鉴定专门参与共代谢生长的基因，我们筛选了脱硫弧菌 G20 的突变体文库，以寻找其与甲烷八叠球菌 JF-1 共代谢生长能力丧失的突变体。获得了 20 个共代谢生长能力受损的突变体。所有 20 个突变体在硫化物条件下以乳酸为唯一碳源纯培养时，生长速度和最大 OD(600)与亲本菌株相似。影响乳酸共代谢生长的突变数量最多的是编码参与 H(2)氧化、电子传递、氢化酶翻译后修饰、丙酮酸降解和信号转导的蛋白质的基因。编码醌还原酶复合物 (Qrc) 的 qrcB 基因和编码周质四血红细胞色素 c(3) (TpIc(3))的 cycA 基因是 G20 与乳酸共代谢生长所必需的。编码 Fe 单氢化酶 (Hyd) 的 hydA 基因的突变体也不能与乳酸共代谢生长。其他突变体在与 M. hungatei JF-1 的共培养中比亲本菌株生长缓慢。qrcB 和 cycA 先前被证明是 G20 纯培养物利用 H(2)和硫酸盐生长所必需的。亲本菌株的洗涤细胞可以从乳酸或丙酮酸中产生 H(2)，但 qrcB、cycA 和 hydA 突变体的洗涤细胞从丙酮酸中产生 H(2)的速度与亲本菌株相似，并且不能从乳酸中产生大量 H(2)。实时定量 PCR 检测显示，与利用乳酸和硫酸盐进行纯培养生长相比，上述三个基因在共代谢生长过程中的表达增加。我们的工作表明，Hyd、Qrc 和 TpIc(3) 参与了 D. alaskensis G20 共代谢乳酸代谢过程中的 H(2)产生，强调了 H(2)产生对该菌株共代谢乳酸代谢的重要性。
