APPLIED GENETICS AND MOLECULAR BIOTECHNOLOGY Proline reduces the binding of transcriptional regulator ArgR to upstream of argB in Corynebacterium glutamicum Soo Youn Lee & Hwa Sung Shin & Jin-Soo Park & Yang-Hoon Kim & Jiho Min Received: 17 August 2009 / Revised: 11 September 2009 / Accepted: 14 September 2009 / Published online: 2 October 2009 # Springer-Verlag 2009 Abstract In this study, the ArgR-binding sites on the arg operon Corynbebacterium glutamicum were characterized by in vivo chromatin immunoprecipitation (ChIP). In addition, the ArgR-binding affinity in the presence of glutamate, proline, or arginine was examined to get further information on expression control. The ChIP assay showed that the ArgR protein binds specifically to the upstream regions of argC, argB, argF , and argG. Upon proline supplementation, ArgR-binding affinity was significantly reduced upstream of argB, resulting in increased ornithine production. In contrast, there was no change in the binding affinity of ArgR to the upstream regions of argC, argF , argG, or argB following the addition of glutamate and arginine. These results suggest that the upstream region of argB on the arg operon plays an important role in interacting with ArgR under proline-supplemented condi- tions and that proline causes an increase in the endogenous level of ornithine by reducing the binding affinity of ArgR to the upstream region of argB. Keywords Proline . Upstream of argB . Corynebacterium glutamicum . ChIP assay . Ornithine Introduction Corynebacterium glutamicum is a gram-positive soil bacterium that is widely used in the industrial production of many amino acids (Leuchtenberger et al. 2005). Ornithine, an intermediate metabolite in arginine biosyn- thesis, is synthesized in C. glutamicum by converting glutamate, through a series of acetylated intermediates, to ornithine in five steps (Lee et al. 2000). The genes involved in the arginine biosynthesis pathway in C. glutamicum are organized as two separate parts, which are argCJBDFR and argGH (Hänßler et al. 2007; Larsen et al. 2005), that encode all of the enzymes required to convert glutamate to arginine via ornithine (Xu et al. 2007; Sakanyan et al. 1996). In this pathway, feedback inhibition control occurs by a single enzyme (N-acetylglutamate kinase encoded by argB), and the expression of the arg operon is not influenced by arginine (Udaka 1966; Sakanyan et al. 1996). Regulation of the arginine biosynthesis pathway has been investigated in Escherichia coli and Bacillus stear- Electronic supplementary material The online version of this article (doi:10.1007/s00253-009-2264-5) contains supplementary material, which is available to authorized users. S. Y. Lee : J. Min Department of Bioprocess Engineering, Chonbuk National University, 664-14 Duckjin-dong, Jeonju 561-756, South Korea H. S. Shin Department of Biological Engineering, Inha University, Incheon 402-751, South Korea J.-S. Park Department of Environmental Engineering, College of Engineering, Sangmyung University, 300 Anseo-dong, Dongnam-gu, Cheonan, Chungnam Province 330-720, South Korea Y.-H. Kim (*) School of Life Science, Chungbuk National University, 410 Sungbong-Ro, Heungduk-Gu, Cheongju 361-763, South Korea e-mail: kyh@chungbuk.ac.kr J. Min (*) Division of Chemical Engineering, Chonbuk National University, 664-14 Duckjin-dong, Jeonju 561-756, South Korea e-mail: jihomin@chonbuk.ac.kr Appl Microbiol Biotechnol (2010) 86:235–242 DOI 10.1007/s00253-009-2264-5