Roles of glucitol in the GutR-mediated transcription activation process in Bacillus subtilis: glucitol induces GutR to change its conformation and to bind ATP.
GutR is a 95-kDa glucitol-dependent transcription activator that mediates the expression of the Bacillus subtilis glucitol operon. Glucitol allows GutR to bind tightly to its binding site located upstream of the gut promoter. In this study, a second functional role of glucitol is identified. Glucitol induces GutR to change its conformation and triggers GutR to bind ATP efficiently. After sequential binding of glucitol and ATP to GutR, GutR adopts a new conformation by forming a compact structure that is resistant to trypsin digestion. Under this condition, the ATP.glucitiol.GutR complex can dissociate slowly from the gutR-binding site (t(12) = 274 min). Interestingly, if ATP in the ATP.glucitiol.GutR complex is replaced by ADP, GutR adopts another conformation and can dissociate from the gutR-binding site even faster (t(12) = 82 min). In all these GutR-DNA binding studies in the presence of different ligands (glucitol, ATP, or ADP), only the off-rate is affected. The vital role of ATP in the GutR-mediated transcription activation process is reflected by the poor transcription from the gut promoter with GutR(D285A) which has a mutation in the motif B of the putative ATP-binding site. A working model for this transcription activation process is presented.
Poon
K K
KK
Division of Cellular, Molecular and Microbial Biology, Department of Biological Sciences, University of Calgary, Calgary, Alberta T2N 1N4, Canada.
Chu
J C
JC
Wong
S L
SL
D000255:Adenosine Triphosphate / Q000378:metabolism; D001412:Bacillus subtilis / Q000235:genetics* / Q000378:metabolism*; D001426:Bacterial Proteins; D001665:Binding Sites; D015153:Blotting, Western; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004926:Escherichia coli / Q000378:metabolism; D007700:Kinetics; D008024:Ligands; D008954:Models, Biological; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010957:Plasmids / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D013012:Sorbitol / Q000494:pharmacology*; D020349:Surface Plasmon Resonance; D013997:Time Factors; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014357:Trypsin / Q000494:pharmacology; D014466:Ultraviolet Rays
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; C087480:GutR protein, Bacillus subtilis; D008024:Ligands; D013012:Sorbitol; D000255:Adenosine Triphosphate; D014357:Trypsin