Cantera J Jason L, Stein Lisa Y
Department of Environmental Sciences, Geology 2207, University of California, Riverside, CA 92521, USA.
Environ Microbiol. 2007 Mar;9(3):765-76. doi: 10.1111/j.1462-2920.2006.01198.x.
Sequences of copper-containing nitrite reductase (nirK) genes obtained from completed nitrifier genome sequences were used to design polymerase chain reaction (PCR) primers to amplify partial nirK sequences from one Nitrosomonas and four Nitrosospira isolates. Deduced NirK protein sequences were highly similar to other copper-containing nitrite reductases including conserved motifs. Phylogenetic comparisons of NirK protein sequences placed orthologues from Nitrosomonas, Nitrosospira and Nitrobacter species into multiple distinct clades. Products related to nirK genes were not amplified from seven additional Nitrosomonas and Nitrosospira isolates by PCR with nirK-specific primers; however, DNA extracted from four of these isolates produced detectable signals in low-stringency Southern hybridizations probed with nirK gene fragments from ammonia-oxidizers with known nirK gene sequences. Analysis of promoter regions of nitrifier nirK genes revealed conserved binding motifs for the NsrR transcription factor in only one clade; other available nitrifier nirK gene promoters lacked characterized transcription factor binding motifs. Taken together, these results indicate that the sequences and regulation of nirK genes are diverse among nitrifiers. This study provides insight to the use of nirK genes for molecular diversity studies, establishes a framework to resolve the origins and diversification of nitrite reduction among the nitrifiers, and expands the database of nirK orthologues.
从已完成的硝化细菌基因组序列中获得的含铜亚硝酸还原酶（nirK）基因序列，被用于设计聚合酶链反应（PCR）引物，以扩增来自一株亚硝化单胞菌和四株亚硝化螺菌分离株的部分nirK序列。推导的NirK蛋白序列与其他含铜亚硝酸还原酶高度相似，包括保守基序。NirK蛋白序列的系统发育比较将来自亚硝化单胞菌、亚硝化螺菌和硝化杆菌物种的直系同源物归入多个不同的进化枝。使用nirK特异性引物通过PCR未从另外七株亚硝化单胞菌和亚硝化螺菌分离株中扩增出与nirK基因相关的产物；然而，从其中四株分离株中提取的DNA在用来自具有已知nirK基因序列的氨氧化菌的nirK基因片段进行低严谨度Southern杂交时产生了可检测信号。对硝化细菌nirK基因启动子区域的分析表明，仅在一个进化枝中存在NsrR转录因子的保守结合基序；其他可用的硝化细菌nirK基因启动子缺乏特征性的转录因子结合基序。综上所述，这些结果表明nirK基因的序列和调控在硝化细菌中是多样的。本研究为nirK基因用于分子多样性研究提供了见解，建立了一个框架来解析硝化细菌中亚硝酸盐还原的起源和多样化，并扩展了nirK直系同源物的数据库。
