Liu Siqing, Dien Bruce S, Cotta Michael A
Bioproducts and Biocatalysis Research Unit, National Center for Agriculture Utilization Research, USDA, ARS, 1815 N. University St., Peoria, IL, 61604, USA.
Curr Microbiol. 2005 Jun;50(6):324-8. doi: 10.1007/s00284-005-4485-x. Epub 2005 Jun 13.
A pyruvate decarboxylase (PDC) gene from bacterial Zymobacter palmae (Zymopdc) was cloned, characterized, and introduced into Lactococcus lactis via a shuttle vector pAK80 as part of a research strategy to develop an efficient ethanol-producing lactic acid bacteria (LAB). The expression levels of Zymopdc gene in the host, as measured by a colorimetric assay based on PDC catalyzed formation of (R)-phenylacetylcarbinol ((R)-PAC), appeared to be dependent on the strength of corresponding Gram-positive promoters. A constitutive, highly expressed promoter conferred the greatest PDC activity, and an acid-inducible promoter demonstrated acid-inducible expression. The metabolic production of ethanol and other products was examined in flask fermentations. More than eightfold increases in acetaldehyde concentrations were detected in two recombinant strains. However, no detectable differences for ethanol fermentation in these engineered strains were observed compared with that of the strain carrying lacZ reporter.
克隆并鉴定了来自巴氏发酵单胞菌（Zymobacter palmae）的丙酮酸脱羧酶（PDC）基因（Zymopdc），并通过穿梭载体pAK80将其导入乳酸乳球菌（Lactococcus lactis），这是开发高效产乙醇乳酸菌（LAB）研究策略的一部分。基于PDC催化生成（R）-苯乙酰甲醇（（R）-PAC）的比色测定法测量，Zymopdc基因在宿主中的表达水平似乎取决于相应革兰氏阳性启动子的强度。组成型高表达启动子赋予最大的PDC活性，酸诱导型启动子表现出酸诱导表达。在摇瓶发酵中检测了乙醇和其他产物的代谢产生。在两个重组菌株中检测到乙醛浓度增加了八倍以上。然而，与携带lacZ报告基因的菌株相比，在这些工程菌株中未观察到乙醇发酵的可检测差异。
