Agrobacterium tumefaciens strain SCUEC1 is a nicotine-degrading bacterium, which has been recently isolated from the tobacco waste-contaminated field soil. However, the mechanism for nicotine degradation in this strain remains unclear. Here, we analyze the function and biological properties of the agnH gene in the strain SCUEC1. The overexpression of the AgnH protein was detected by SDS-PAGE analysis, and functional insight of the AgnH protein was carried out with monitoring the changes of maleic acid into fumaric acid by high performance liquid chromatography (HPLC). Moreover, the effects of temperature, pH and metal ions on the enzymatic activities of the AgnH protein were also analyzed. The results demonstrated that the agnH gene was successfully ligated to the plasmid pET28a. The optimal condition for the enzymatic activities for the AgnH, approximately 28.0 kDa, was determined as 37 °C, pH 8.0, and 25 µM Mg2+. Conclusively, the agnH gene fulfils an important role in the conversion of maleic acid into fumaric acid involved in nicotine-degradation pathways in Agrobacterium tumefaciens strain SCUEC1.

中文翻译：

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根癌农杆菌菌株SCUEC1中涉及烟碱降解途径的agnH基因的功能分析。

根癌农杆菌菌株SCUEC1是降解尼古丁的细菌，最近已从被烟草废料污染的田间土壤中分离出来。但是，该菌株中尼古丁降解的机制仍不清楚。在这里，我们分析了菌株SCUEC1中agnH基因的功能和生物学特性。通过SDS-PAGE分析检测到AgnH蛋白的过表达，并通过高效液相色谱法（HPLC）监测马来酸向富马酸的变化来进行AgnH蛋白的功能研究。此外，还分析了温度，pH和金属离子对AgnH蛋白酶活性的影响。结果表明，agnH基因已成功连接到质粒pET28a。AgnH的酶促活性的最佳条件，在37°C，pH 8.0和25 µM Mg2 +的条件下测定大约28.0 kDa。结论是，agnH基因在根癌农杆菌菌株SCUEC1的顺丁烯二酸降解途径中的马来酸转化为富马酸中起重要作用。