Cloning, sequencing, and functional expression in Escherichia coli of chaperonin (groESL) genes from Vibrio cholerae.
Using a series of oligonucleotides synthesized on the basis of conserved nucleotide motifs in heat-shock genes, the groESL heat-shock operon from a Vibrio cholerae TSI-4 strain has been cloned and sequenced, revealing that the presence of two open reading frames (ORFs) of 291 nucleotides and 1,632 nucleotides separated by 54 nucleotides. The first ORF encoded a polypeptide of 97 amino acids, GroES homologue, and the second ORF encoded a polypeptide of 544 amino acids, GroEL homologue. A comparison of the deduced amino acid sequences revealed that the primary structures of the V. cholerae GroES and GroEL proteins showed significant homology with those of the GroES and GroEL proteins of other bacteria. Complementation experiments were performed using Escherichia coli groE mutants which have the temperature-sensitive growth phenotype. The results showed that the groES and groEL from V. cholerae were expressed in E. coli, and groE mutants harboring V. cholerae groESL genes regained growth ability at high temperature. The evolutionary analysis indicates a closer relationship between V. cholerae chaperonins and those of the Haemophilus and Yersinia species.
Mizunoe
Y
Y
Department of Bacteriology, Faculty of Medicine, Kyushu University, Fukuoka, Japan. ymizunoe@bact.med.kyushu-u.ac.jp
Wai
S N
SN
Umene
K
K
Kokubo
T
T
Kawabata
S
S
Yoshida
S
S
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000502:physiology; D001483:Base Sequence; D018835:Chaperonin 10 / Q000737:chemistry / Q000235:genetics / Q000502:physiology; D018834:Chaperonin 60 / Q000737:chemistry / Q000235:genetics / Q000502:physiology; D018833:Chaperonins / Q000737:chemistry / Q000235:genetics* / Q000502:physiology; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D005798:Genes, Bacterial; D005816:Genetic Complementation Test; D018869:Heat-Shock Response; D008969:Molecular Sequence Data; D010802:Phylogeny; D011994:Recombinant Proteins; D017422:Sequence Analysis, DNA; D014734:Vibrio cholerae / Q000235:genetics*
D001426:Bacterial Proteins; D018835:Chaperonin 10; D018834:Chaperonin 60; C097970:GroESL protein, Bacteria; D011994:Recombinant Proteins; D018833:Chaperonins