Sequence analysis and heterologous expression of the groE genes from Thermoanaerobacter sp. Rt8.G4.
The groE homologous genes of the anaerobic thermophile Thermoanaerobacter sp. Rt8.G4 (TRt) have been isolated, sequenced and analysed. The TRt groES and groEL encode subunits of chaperonin 10 (Cpn10) and chaperonin 60 (Cpn60) of 94 and 541 amino acids, respectively, and are arranged in that order forming the open reading frames (ORFs) of a bicistronic operon. A controlling inverted repeat of chaperone expression (CIRCE) element lies between the consensus promoter of the operon and TRt groES. At optimum growth temperature (65 degreesC) the chaperonins of TRt are expressed, but production of Cpn60 increases significantly following temperature increases of 3-10 degreesC. Functionally intact recombinant TRt chaperonins were produced in Escherichia coli. However, owing to codon incompatibility, replacement of consecutive AGA codons in the gene encoding TRt Cpn60 was necessary for optimum expression in this heterologous host.
Truscott
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School of Biochemistry, La Trobe University, Bundoora, Victoria, 3083, Australia.
Scopes
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RK
D000595:Amino Acid Sequence; D001421:Bacteria, Anaerobic / Q000235:genetics*; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics*; D018835:Chaperonin 10 / Q000235:genetics; D018834:Chaperonin 60 / Q000235:genetics; D018833:Chaperonins; D016384:Consensus Sequence; D017931:DNA Primers; D004926:Escherichia coli / Q000235:genetics; D029968:Escherichia coli Proteins; D005798:Genes, Bacterial; D016988:Gram-Positive Asporogenous Rods, Irregular / Q000235:genetics*; D006360:Heat-Shock Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics*; D006358:Hot Temperature; D016297:Mutagenesis, Site-Directed; D016366:Open Reading Frames; D009876:Operon; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry
D001426:Bacterial Proteins; D018835:Chaperonin 10; D018834:Chaperonin 60; D017931:DNA Primers; D029968:Escherichia coli Proteins; C028715:GroE protein, E coli; D006360:Heat-Shock Proteins; D011994:Recombinant Proteins; D018833:Chaperonins