SEQUENCE-ANALYSIS AND HETEROLOGOUS EXPRESSION OF THE GROE GENES FROM THERMOANAEROBACTER SP. RT8.G4

The groE homologous genes of the anaerobic thermophile Thermoanaerobac ter sp. Rt8.G4 (TRt) have been isolated, sequenced and analysed. The T Rt groES and groEL encode subunits of chaperonin 10 (Cpn10) and chaper onin 60 (Cpn60) of 94 and 541 amino acids, respectively, and are arran ged in that order forming the open reading frames (ORFs) of a bicistro nic operon. A controlling inverted repeat of chaperone expression (CIR CE) element lies between the consensus promoter of the operon and TRt groES. At optimum growth temperature (65 degrees C) the chaperonins of TRt are expressed, but production of Cpn60 increases significantly fo llowing temperature increases of 3-10 degrees C. Functionally intact r ecombinant 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 i n this heterologous host. (C) 1998 Elsevier Science B.V. All rights re served.