Nucleotide sequences of genes encoding heat-stable and heat-labile glyceraldehyde-3-phosphate dehydrogenases; amino acid sequence and protein thermostability.
The structural gene (gapST) encoding glyceraldehyde-3-phosphate dehydrogenase (GPDH; EC 1.2.1.12) from Bacillus stearothermophilus has been cloned in Escherichia coli using plasmid pBR322 as a vector; the homologous gene (gapCO) from Bacillus coagulans was cloned from a phage lambda library. Expression of the cloned gap genes revealed that, as in the wild-type (wt) organisms, the GPDH from B. stearothermophilus (GPDH-ST) was intrinsically heat stable (hs) and that from B. coagulans (GPDH-CO) heat labile (hl). The cloned gap genes were sequenced and the deduced amino acid (aa) sequences were found to be highly conserved (91.6% homology), despite the large difference in thermostability between these two enzymes. Of the 28 aa which differ between the two proteins, most of which occur in the middle third of the monomeric subunit, 5 aa involve replacement of alanine in the hl GPDH-CO, by proline in the hs GPDH-ST, and are especially interesting in terms of their potential contributions to thermostability. Conservation at the DNA level is equally dramatic, with the two gap genes exhibiting 93.3% nucleotide sequence homology. These highly expressed genes exhibit an equivalent codon bias, which more closely resembles that of highly expressed E. coli genes, than that of B. stearothermophilus genes whether highly or weakly expressed.
Tesfay
H S
HS
Department of Microbiology, Molecular Genetics and Immunology, University of Kansas Medical Center, College of Health Sciences and Hospital, Kansas City 66103.
Amelunxen
R E
RE
Goldberg
I D
ID
D000595:Amino Acid Sequence; D001407:Bacillus / Q000235:genetics*; D001426:Bacterial Proteins / Q000235:genetics*; D003062:Codon; D004926:Escherichia coli; D005798:Genes, Bacterial; D001411:Geobacillus stearothermophilus / Q000235:genetics; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000235:genetics*; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000235:genetics; D012689:Sequence Homology, Nucleic Acid; D013045:Species Specificity
D001426:Bacterial Proteins; D003062:Codon; D011993:Recombinant Fusion Proteins; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases