Wang H, Tseng C P, Gunsalus R P
Department of Microbiology and Molecular Genetics, University of California, Los Angeles, California 90095-1489, USA.
J Bacteriol. 1999 Sep;181(17):5303-8. doi: 10.1128/JB.181.17.5303-5308.1999.
Escherichia coli synthesizes two biochemically distinct nitrate reductase enzymes, a membrane-bound enzyme encoded by the narGHJI operon and a periplasmic cytochrome c-linked nitrate reductase encoded by the napFDAGHBC operon. To address why the cell makes these two enzymes, continuous cell culture techniques were used to examine napF and narG gene expression in response to different concentrations of nitrate and/or nitrite. Expression of the napF-lacZ and narG-lacZ reporter fusions in strains grown at different steady-state levels of nitrate revealed that the two nitrate reductase operons are differentially expressed in a complementary pattern. The napF operon apparently encodes a "low-substrate-induced" reductase that is maximally expressed only at low levels of nitrate. Expression is suppressed under high-nitrate conditions. In contrast, the narGHJI operon is only weakly expressed at low nitrate levels but is maximally expressed when nitrate is elevated. The narGHJI operon is therefore a "high-substrate-induced" operon that somehow provides a second and distinct role in nitrate metabolism by the cell. Interestingly, nitrite, the end product of each enzyme, had only a minor effect on the expression of either operon. Finally, nitrate, but not nitrite, was essential for repression of napF gene expression. These studies reveal that nitrate rather than nitrite is the primary signal that controls the expression of these two nitrate reductase operons in a differential and complementary fashion. In light of these findings, prior models for the roles of nitrate and nitrite in control of narG and napF expression must be reconsidered.
大肠杆菌合成两种生化特性不同的硝酸还原酶，一种是由narGHJI操纵子编码的膜结合酶，另一种是由napFDAGHBC操纵子编码的周质细胞色素c连接的硝酸还原酶。为了探究细胞为何产生这两种酶，采用连续细胞培养技术来检测napF和narG基因对不同浓度硝酸盐和/或亚硝酸盐的表达响应。在不同稳态硝酸盐水平下生长的菌株中，napF-lacZ和narG-lacZ报告基因融合体的表达表明，这两个硝酸还原酶操纵子以互补模式差异表达。napF操纵子显然编码一种“低底物诱导”的还原酶，仅在低硝酸盐水平时才最大程度表达。在高硝酸盐条件下表达受到抑制。相反，narGHJI操纵子在低硝酸盐水平时仅微弱表达，但在硝酸盐水平升高时最大程度表达。因此，narGHJI操纵子是一种“高底物诱导”操纵子，它以某种方式在细胞的硝酸盐代谢中发挥第二种不同的作用。有趣的是，每种酶的终产物亚硝酸盐对任一操纵子的表达影响都很小。最后，硝酸盐而非亚硝酸盐对于napF基因表达的抑制至关重要。这些研究表明，硝酸盐而非亚硝酸盐是主要信号，以差异和互补的方式控制这两个硝酸还原酶操纵子的表达。鉴于这些发现，必须重新考虑先前关于硝酸盐和亚硝酸盐在控制narG和napF表达中作用的模型。
