Promoter/repressor system of Lactobacillus plantarum phage og1e: characterization of the promoters pR49-pR-pL and overproduction of the cro-like protein cng in Escherichia coli.
The Lactobacillus plantarum phage og1e (42<HSP SP = "0. 25">259<HSP SP = "0.25">bp) has two repressor-like genes cng and cpg oriented oppositely, accompanied by three potential promoters pR, pL and pR49, and seven operator-like sequences (GATAC-boxes) (Kodaira et al., 1997). In this study, the og1e putative promoters were introduced into the Escherichia coli promoter-detecting plasmid pKK232-8. In E. coli CK111, pR (pKPR1), pL (pKPL1) and pR49 (pKPR49) exhibited distinct CAT activities. When pKPR1 or pKPL1 was coexistent with a compatible plasmid pACYC184 carrying pR-cng (pA4PRCN1), the CAT activity was decreased significantly. On the other hand, cng directed a protein (Cng) of 10.1 kDa in E. coli under the control of T7 promoter. Gel mobility-shift assays demonstrated that Cng binds specifically to a DNA region containing the GATAC-boxes. In addition, primer extension analyses demonstrated that the two sequences pR and pL act as a promoter in L. plantarum as well as in E. coli. These results suggested that the potential promoters pR and pL probably function for the lytic and lysogenic pathways, respectively, and Cng may act as a repressor presumably through the GATAC-boxes as operators.
Molecular Biology Group, Faculty of Engineering, Toyama University, 3190, Gofuku, Toyama 930, Japan.
M
M
Kakikawa
N
N
Watanabe
T
T
Funawatashi
M
M
Oki
H
H
Yasukawa
A
A
Taketo
K I
KI
Kodaira
D001435:Bacteriophages / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites; D015500:Chloramphenicol O-Acetyltransferase / Q000096:biosynthesis; D003001:Cloning, Molecular; D004268:DNA-Binding Proteins; D004926:Escherichia coli / Q000235:genetics; D005814:Genes, Viral; D005838:Genotype; D007778:Lactobacillus / Q000235:genetics / Q000821:virology*; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D010957:Plasmids; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000378:metabolism; D012097:Repressor Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D017361:Viral Nonstructural Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D014764:Viral Proteins; D054334:Viral Regulatory and Accessory Proteins; D015678:Viral Structural Proteins / Q000235:genetics*
C114486:Cng protein, Bacteriophage phig1e; D004268:DNA-Binding Proteins; D009838:Oligodeoxyribonucleotides; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D017361:Viral Nonstructural Proteins; D014764:Viral Proteins; D054334:Viral Regulatory and Accessory Proteins; D015678:Viral Structural Proteins; C016649:phage repressor proteins; D015500:Chloramphenicol O-Acetyltransferase