Nitrogen metabolism in Streptomyces coelicolor A3(2):: modification of glutamine synthetase I by an adenylyltransferase
Nitrogen metabolism in Streptomyces coelicolor A3(2):: modification of glutamine synthetase I by an adenylyltransferase
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DOI：
10.1099/00221287-145-9-2313
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发表时间：
1999-09-01
期刊：
影响因子：
--
通讯作者：
Engels, A
中科院分区：
文献类型：
--
作者：
Fink, D;Falke, D;Engels, A
利用基于共识基序的异源PCR引物，扩增了链霉菌内部腺苷基转移酶基因（glnE）片段。该序列与细菌glnE基因具有显著的相似性，并且包含glnE c端腺苷基转移酶结构域的典型基序。来自colcololor的glnE位于染色体的Asel-C片段上，定位在glnA（编码谷氨酰胺合成酶I， GSI）和glnII（编码GSII）附近。为了分析GlnE在葡萄球菌中的功能，构建了GlnE（葡萄球菌E4）和glnA（葡萄球菌HT107）基因替代突变体。氨休克后glnE突变体的GSI活性没有下调。然而，野生型细胞的GSI活性下降到原始活性的60%。glnA突变体不是谷氨酰胺营养不良，但在g-谷氨酰转移酶试验中，未移位和移位的HT107细胞未检测到GSI活性。经蛇毒磷酸二酯酶处理后，野生型的GSI活性得以重建，而E4突变体的GSI活性未见改变。此外，在E4突变体中，短期GSI调节的丧失伴随着谷氨酰胺：谷氨酸比值的增加。
An internal adenylyltransferase gene (glnE) fragment from Streptomyces coelicolor was amplified using heterologous PCR primers derived from consensus motifs. The sequence had significant similarity to bacterial glnE genes, and included a motif typical of the C-terminal adenylyltransferase domain of GlnE. glnE from S. coelicolor lies on the Asel-C fragment of the chromosome and is localized near glnA (encoding glutamine synthetase I, GSI) and glnII (encoding GSII). To analyse the function of GlnE in S. coelicolor, glnE (S. coelicolor E4) and glnA (S. coelicolor HT107) gene replacement mutants were constructed. The GSI activity of the glnE mutant was not down-regulated after an ammonium shock. However, the GSI activity of the wild-type cells decreased to 60% of the original activity. The glnA mutant is not glutamine auxotrophic, but in the g-glutamyltransferase assay no GSI activity was detected in unshifted and shifted HT107 cells. By snake venom phosphodiesterase treatment the GSI activity in the wild-type can be reconstituted, whereas no alteration is observed in the E4 mutant. Additionally, the loss of short-term GSI regulation in the E4 mutant was accompanied by an increased glutamine:glutamate ratio.