Expression and Characterization of ArgR, An Arginine Regulatory Protein in Corynebacterium crenatum

• Published in: Biomedical and environmental sciences 2014-06, Vol.27 (6), p.436-443
• Main Authors: CHEN, Xue Lan, ZHANG, Bin, TANG, Li, JIAO, Hai Tao, XU, Heng Yi, XU, Feng, XU, Hong, WEI, Hua, XIONG, Yong Hua
• Format: Article
• Language: English
• Subjects: Arginine - biosynthesis; Arginine biosynthetic genes; ArgR protein; Bacterial Proteins - chemistry; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Corynebacterium - genetics; Corynebacterium - metabolism; Corynebacterium crenatum; Electrophoretic mobility shift assay; Gene Expression Regulation, Bacterial; gR基因; Real-time PCR; Repressor Proteins - chemistry; Repressor Proteins - genetics; Repressor Proteins - metabolism; 氨基酸分析仪; 生产检测; 生物合成基因; 精氨酸; 表征; 调节蛋白; 钝齿棒杆菌
• ISSN: 0895-3988; 2214-0190
• DOI: 10.3967/bes2014.072

Objective Corynebacterium crenatum MT, a mutant from C. crenatum AS 1.542 with a lethal argR gene, exhibits high arginine production. To confirm the effect of ArgR on arginine biosynthesis in C. crenatum, an intact argR gene from wild-type AS 1.542 was introduced into C. crenatum MT, resulting in C. crenatum MT. sp, and the changes of transcriptional levels of the arginine biosynthetic genes and arginine production were compared between the mutant strain and the recombinant strain. Methods Quantitative real-time polymerase chain reaction was employed to analyze the changes of the related genes at the transcriptional level, electrophoretic mobility shift assays were used to determine ArgR binding with the argCJBDF, argGH, and carAB promoter regions, and arginine production was determined with an automated amino acid analyzer. Results Arginine production assays showed a 69.9% reduction in arginine from 9.01±0.22 mg/mL in C. crenatum MT to 2.71±0.13 mg/mL （P〈0.05） in C. crenatum MT. sp. The argC, argB, argD, argF, argJ, argG, and carA genes were down-regulated significantly in C. crenatum MT. sp compared with those in its parental C. crenatum MT strain. The electrophoretic mobility shift assays showed that the promoter regions were directly bound to the ArgR protein. Conclusion The arginine biosynthetic genes in C crenatum are clearly controlled by the regulator ArgR, and intact ArgR in C. crenatum MT results in a significant descrease in production. negative arginine production.