title: Purification and characterization of an arginine regulatory protein, ArgR, in Corynebacterium glutamicum.
abstract: Corynebacterium glutamicum, a Gram-positive bacterium, has been widely used for industrial amino acid production. We previously showed that, in C. glutamicum, argCJBDFRGH arginine biosynthesis genes are clustered but independently transcribed from argC and argG promoters, leading to the generation of two transcripts corresponding to argCJBDFR and argGH. In this report, we show the effect of the C. glutamicum ArgR repressor on argC and argG promoters by overexpressing or disrupting the argR gene. Gel filtration assay results indicate that native ArgR is a hexamer of equal subunits with molecular mass of 110 kDa. Protein sequence analysis revealed the presence of an "SR" (Ser₅₇-Arg₅₈) motif for the DNA binding site at the N-terminal region and the "GTIAGDDTV" motif for arginine binding and its oligomerization at the C-terminal region. An argC or argG promoter-lacZ fusion reporter assay and argR mutational analysis showed that transcription of the argCJBDFR arginine biosynthesis genes is regulated from the argC promoter by ArgR in cooperation with L-arginine in C. glutamicum. This finding was supported by the gel mobility-shift assay showing direct binding of hexameric ArgR to the argC promoter in the presence of L-arginine. Unexpectedly, argGH transcription was not responsive to the level of ArgR repressor and/or arginine. In a further study, a C. glutamicum argR mutant was constructed by disrupting the chromosomal argR gene to manufacture an improved arginine-producing strain. Arginine productivity was increased in the C. glutamicum argR mutant strain under conditions of both limited and excessive arginine.
authors.affiliation: Division of Biological Science and Research Center for Women's Diseases, Sookmyung Women's University, Seoul 140-742, South Korea.
authors.forename: Sei-Hyun
authors.initials: SH
authors.lastname: Yim
authors.forename: Samil
authors.initials: S
authors.lastname: Jung
authors.forename: Shun-Kyoung
authors.initials: SK
authors.lastname: Lee
authors.forename: Choong-Il
authors.initials: CI
authors.lastname: Cheon
authors.forename: Eunsook
authors.initials: E
authors.lastname: Song
authors.forename: Soo-Suk
authors.initials: SS
authors.lastname: Lee
authors.forename: Jin
authors.initials: J
authors.lastname: Shin
authors.forename: Myeong-Sok
authors.initials: MS
authors.lastname: Lee
mesh_terms: D000818:Animals; D001120:Arginine / Q000096:biosynthesis*; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D001483:Base Sequence; D048230:Corynebacterium glutamicum / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D012045:Regulatory Sequences, Nucleic Acid; D012097:Repressor Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D016415:Sequence Alignment
chemical_list: C087018:ArgR protein, Bacteria; D001426:Bacterial Proteins; D012097:Repressor Proteins; D001120:Arginine