Li Xiangkai, Krumholz Lee R
Department of Botany and Microbiology, University of Oklahoma, Norman, OK 73019-0245, USA.
J Bacteriol. 2007 May;189(10):3705-11. doi: 10.1128/JB.01913-06. Epub 2007 Mar 2.
Desulfovibrio desulfuricans G20 grows and reduces 20 mM arsenate to arsenite in lactate-sulfate media. Sequence analysis and experimental data show that D. desulfuricans G20 has one copy of arsC and a complete arsRBCC operon in different locations within the genome. Two mutants of strain G20 with defects in arsenate resistance were generated by nitrosoguanidine mutagenesis. The arsRBCC operons were intact in both mutant strains, but each mutant had one point mutation in the single arsC gene. Mutants transformed with either the arsC1 gene or the arsRBCC operon displayed wild-type arsenate resistance, indicating that the two arsC genes were equivalently functional in the sulfate reducer. The arsC1 gene and arsRBCC operon were also cloned into Escherichia coli DH5alpha independently, with either DNA fragment conferring increased arsenate resistance. The recombinant arsRBCC operon allowed growth at up to 50 mM arsenate in LB broth. Quantitative PCR analysis of mRNA products showed that the single arsC1 was constitutively expressed, whereas the operon was under the control of the arsR repressor protein. We suggest a model for arsenate detoxification in which the product of the single arsC1 is first used to reduce arsenate. The arsenite formed is then available to induce the arsRBCC operon for more rapid arsenate detoxification.
脱硫脱硫弧菌G20在乳酸 - 硫酸盐培养基中生长，并将20 mM砷酸盐还原为亚砷酸盐。序列分析和实验数据表明，脱硫脱硫弧菌G20在基因组内的不同位置有一个arsC拷贝和一个完整的arsRBCC操纵子。通过亚硝基胍诱变产生了两株对砷酸盐抗性有缺陷的G20突变株。两个突变株中的arsRBCC操纵子均完整，但每个突变株的单个arsC基因都有一个点突变。用arsC1基因或arsRBCC操纵子转化的突变株表现出野生型砷酸盐抗性，这表明两个arsC基因在硫酸盐还原菌中具有同等功能。arsC1基因和arsRBCC操纵子也分别独立克隆到大肠杆菌DH5α中，两个DNA片段都赋予了增强的砷酸盐抗性。重组的arsRBCC操纵子使菌株能够在LB肉汤中50 mM的砷酸盐浓度下生长。对mRNA产物的定量PCR分析表明，单个arsC1组成型表达，而操纵子受arsR阻遏蛋白的控制。我们提出了一种砷酸盐解毒模型，其中单个arsC1的产物首先用于还原砷酸盐。然后形成的亚砷酸盐可诱导arsRBCC操纵子进行更快的砷酸盐解毒。
