Characterization of glutamine synthetase from the ammonium-excreting strain HM053 of Azospirillum brasilense
Characterization of glutamine synthetase from the ammonium-excreting strain HM053 of Azospirillum brasilense
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DOI：
10.1590/1519-6984.235927
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发表时间：
2022-01-01
影响因子：
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通讯作者：
Souza, Emanuel Maltempi
中科院分区：
文献类型：
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作者：
Ghenov, Fernanda;Gerhardt, Edileusa Cristina Marques;Souza, Emanuel Maltempi
谷氨酰胺合成酶（GS）由glnA编码，催化L-谷氨酸和铵转化为L-谷氨酰胺。这一ATP水解驱动的过程是固氮细菌Azoacellum brasilense的主要氮同化途径。助理巴西根瘤菌HM 053菌株GS活性差，在固氮条件下会向培养基中渗漏铵。本研究将野生型和HM 053菌株的glnA基因克隆到pET 28a中，测序并在E.杆菌GS酶通过亲和层析纯化并表征。HM 053菌株的GS携带P347L取代，这导致酶活性低，并且使酶对腺苷酰转移酶GlnE的腺苷酰化不敏感。
Glutamine synthetase (GS), encoded by glnA, catalyzes the conversion of L-glutamate and ammonium to L-glutamine. This ATP hydrolysis driven process is the main nitrogen assimilation pathway in the nitrogen-fixing bacterium Azospirillum brasilense. The A. brasilense strain HM053 has poor GS activity and leaks ammonium into the medium under nitrogen fixing conditions. In this work, the glnA genes of the wild type and HM053 strains were cloned into pET28a, sequenced and overexpressed in E. coli. The GS enzyme was purified by affinity chromatography and characterized. The GS of HM053 strain carries a P347L substitution, which results in low enzyme activity and rendered the enzyme insensitive to adenylylation by the adenilyltransferase GlnE.