Cloning, characterization and functional analysis of groESL operon from thermophilic cyanobacterium Synechococcus vulcanus.
Genes encoding 10914 Da and 58267 Da polypeptides homologous to groES and groEL of Escherichia coli were cloned and sequenced from a thermophilic cyanobacterium, Synechococcus vulcanus. The deduced amino acid sequence of the GroEL protein was much more homologous to GroELs of other cyanobacteria which accompany GroES than another GroEL homolog of S. vulcanus (GroEL2) reported previously (M. Furuki, N. Tanaka, T. Hiyama, and H. Nakamoto, Biochim. Biophys. Acta 1294 (1996) 106-110). We designate the gene as groEL1 to distinguish it from the non-operon forming groEL2 gene. A 9-base pair inverted repeat sequence (TTAGCACTC-N9-GAGTGCTAA) was located upstream of the promoter region of groEL1, which was absent in groEL2. Southern blot analysis indicated that only one groESL1 operon was present in the genomic DNA of S. vulcanus. The amount of the bicistronic, 2.3 kb transcript of groESL1 operon increased 30-fold within 30 min upon heat shock. The increase was completely inhibited by chloramphenicol, suggesting the involvement of heat-induced production of a polypeptide. Introduction of the cloned groEL1 gene into a groEL defective mutant of E. coli resulted in the complementation of heat sensitivity, which contrasted with the previous result with groEL2.
Tanaka
N
N
Department of Biochemistry and Molecular Biology, Saitama University, Urawa, Japan.
Hiyama
T
T
Nakamoto
H
H
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D015152:Blotting, Northern; D018834:Chaperonin 60 / Q000737:chemistry / Q000235:genetics*; D018833:Chaperonins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D002701:Chloramphenicol / Q000494:pharmacology; D003001:Cloning, Molecular; D000458:Cyanobacteria / Q000235:genetics*; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics / Q000254:growth & development; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D018869:Heat-Shock Response; D008969:Molecular Sequence Data; D009876:Operon; D012333:RNA, Messenger / Q000032:analysis; D015183:Restriction Mapping; D012293:Rifampin / Q000494:pharmacology; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D014158:Transcription, Genetic; D014169:Transformation, Bacterial
D001426:Bacterial Proteins; D018834:Chaperonin 60; C097970:GroESL protein, Bacteria; D012333:RNA, Messenger; D002701:Chloramphenicol; D018833:Chaperonins; D012293:Rifampin