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 nu cleotide motifs in heat-shock genes, the groESL heat-shock operon from a Vi brio cholerae TSI-4 strain has been cloned and sequenced, revealing that th e presence of two open reading frames (ORFs) of 291 nucleotides and 1,632 n ucleotides separated by 54 nucleotides, The first ORF encoded a polypeptide of 97 amino acids, GroES homologue, and the second ORF encoded a polypepti de of 544 amino acids, GroEL homologue, A comparison of the deduced amino a cid sequences revealed that the primary structures of the V. cholerae GroES and GroEL proteins showed significant homology with those of the CroES and GroEL proteins of other bacteria, Complementation experiments were perform ed using Escherichia coli groE mutants which have the temperature-sensitive growth phenotype, The results showed that the groES and groEL from V. chol erae were expressed in E. coli, and groE mutants harboring V. cholerae groE SL genes regained growth ability at high temperature, The evolutionary anal ysis indicates a closer relationship between V. cholerae chaperonins and th ose of the Haemophilus and Yersinia species.