Identification and analysis of the dissimilatory nitrous oxide reduction genes, nosRZDFY, of Rhizobium meliloti
Identification and analysis of the dissimilatory nitrous oxide reduction genes, nosRZDFY, of Rhizobium meliloti
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DOI：
10.1128/jb.178.6.1505-1514.1996
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发表时间：
1996-03-01
影响因子：
3.2
通讯作者：
Chan, YK
中科院分区：
文献类型：
--
作者：
Holloway, P;McCormick, W;Chan, YK
在带有nod大质粒的10.1-kb EcoRI片段的粘粒（pYC 7）中鉴定了内共生固氮菌苜蓿根瘤菌（Rhizobium meliloti）异化一氧化氮还原所必需的完整nos基因区。通过Southern杂交和Tn 5诱变将该基因区定位于fixGHIS簇下游8 kb内，一个4.6 kb的DNA片段，包括结构基因nosZ及其侧翼区的核苷酸序列测定显示序列同源性和相似性的遗传组织与斯氏假单胞菌Zobell的nosRZDFY基因。基因被排列成三个互补组，包括nosZ结构基因、nosR调节基因和nosDFY铜加工基因。苜蓿nosZ产物（一种多铜一氧化二氮还原酶）与副球菌属和假单胞菌属的类似基因产物的相似性高于真养产碱菌。nosZ基因之前是nosR，其编码含有C-末端半胱氨酸簇的调节蛋白，所述半胱氨酸簇类似于存在于细菌铁氧还蛋白的4Fe-4S结合区中的那些。nosDFY基因位于nosZ基因下游，编码周质蛋白、ATP/GTP结合蛋白和膜蛋白，可能构成一个铜加工系统。苜蓿草。在nos基因之前没有检测到rpoN结合位点，并且在nos基因区域中的Tn 5插入没有影响共生固氮能力。
The complete nos region essential for dissimilatory nitrous oxide reduction by the endosymbiotic diazotroph Rhizobium meliloti was identified in a cosmid (pYC7) carrying a 10.1-kb EcoRI fragment of the nod megaplasmid, This gene region was localized by Southern hybridization and Tn5 mutagenesis to within 8 kb downstream from the fixGHIS cluster, Nucleotide sequence determination of a 4.6-kb DNA segment including the structural gene nosZ and its flanking regions showed sequence homology and similarity in genetic organization with the nosRZDFY genes of Pseudomonas stutzeri Zobell. The genes were arranged in three complementation groups, comprising the nosZ structural gene, the nosR regulatory gene, and the nosDFY copper-processing genes, The derived amino acid sequence of the R. meliloti nosZ product (a multi-copper nitrous oxide reductase) was more similar to those of the analogous gene products of Paracoccus and Pseudomonas species than to that of Alcaligenes eutrophus. The nosZ gene was preceded by nosR, which encodes a regulatory protein containing C-terminal cysteine clusters similar to those present in the 4Fe-4S binding region of bacterial ferredoxins. The nosDFY genes, located downstream from nosZ, were identified as copper-processing genes encoding a periplasmic protein, an ATP/GTP-binding protein, and a membrane protein, presumably forming a copper-processing system, A consensus sequence for an Anr- or Fnr-binding site similar to that in the upstream sequence of nosZ in Paracoccus denitrificans or P. stutzeri was absent in R. meliloti. No rpoN-binding site preceding the nos genes was detected, and none of the Tn5 insertions in the nos gene region affected symbiotic N-2-fixing ability.