Transketolase A, an enzyme in central metabolism, derepresses the marRAB multiple antibiotic resistance operon of Escherichia coli by interaction with MarR.
The Escherichia coli marRAB operon specifies two regulatory proteins, MarR (which represses) and MarA (which activates expression of the operon). The latter controls expression of multiple other chromosomal genes implicated in cell physiology, multiple drug resistance and virulence. Using randomly cloned E. coli DNA fragments in the bacterial adenylate cyclase two-hybrid system, we found that transketolase A (TktA) interacts with MarR. Purified (6H)-TktA immobilized on NiNTA resin-bound MarR. Overexpression or deletion of tktA showed that TktA interfered with MarR repression of the marRAB operon. Deletion of tktA increased antibiotic and oxidative stress susceptibilities, while its overexpression decreased them. Hydrogen peroxide induced tktA at 1 h treatment, while an increase in marRAB expression occurred only after 3 h exposure. This increase was dependent on the presence of tktA. Two MarR mutations which eliminated MarR binding to the marRAB operator and one which decreased dimerization of MarR had no effect on MarR interaction with TktA in the two-hybrid system. However, the interaction was disrupted by one of the three tested superrepressor mutant MarR proteins known to increase MarR binding to DNA. TktA inhibition of repression by MarR demonstrates a previously unrecognized level of control of the expression of marRAB operon.
Domain
Francis
F
Center for Adaptation Genetics and Drug Resistance, Department of Molecular Biology and Microbiology, Tufts University School of Medicine, Boston, MA 02111, USA.
Bina
Xiaowen R
XR
Levy
Stuart B
SB
D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019281:Dimerization; D024901:Drug Resistance, Multiple, Bacterial / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D009154:Mutation; D009876:Operon; D011485:Protein Binding; D012097:Repressor Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014174:Transketolase / Q000235:genetics / Q000378:metabolism*; D020798:Two-Hybrid System Techniques
D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C080002:MarA protein, E coli; C080004:MarR protein, E coli; D012097:Repressor Proteins; D014174:Transketolase