Characterization of genes associated with glyoxal resistance in escherichia coli : yqhCD and nemRAgloA operon

Abstract

The reactive α-oxoaldehydes such as glyoxal (GO) and methylglyoxal (MG) are generated in vivo from sugars through oxdative stress. GO and MG are believed to be removed from cells by glutathione-dependent glyoxalases and other aldehyde reductases. We isolated a number of GO-resistant mutants from the E. coli strain MG1655 on LB plates containing 10 mM GO. By tagging the mutations with the transposon TnphoA-132 and by determining their co-transductional linkages, we were able to identify two loci where GO resistance $(GO^R)$ mutations were mapped. DNA sequencing of the loci revealed that one of the loci is yqhC gene, which is predicted to encode an AraC-type transcriptional regulator of unknown function, and the other is nemRA operon, which were previously reported to encode a repressor and the N-ethylmaleimide reductase, respectively. In partⅠ, the GO missense mutations were found in predicted regulatory region of YqhC protein, thereby constitutively activating the product of the adjacent gene yqhD. Functions of YqhC and YqhD were further characterized as a transcriptional activator of yqhD that is inducible by GO and as a NADPH-dependent aldehyde reductase that converts GO to 1,2-ethandiol via glycolaldehyde, respectively. In partⅡ, the GO resistance mutations are located in nemR binding site in 5’UTR of nemRA opreon or inframe region of nemR. The mutations enhance both nemA and gloA, encoding glyoxalaseⅠ located downstream of nemA. It was demonstrated that the GO resistance is dependent on gloA but not on nemA, and gloA is expressed as part of the nemRA operon. In addition, it was shown that NemA catalyzes quinone, playing on important role in electron transfer and redox regulation, to quinol using NADPH. GO and quinone dissociates NemR from DNA and derepress nemRA-gloA operon. Intracellular ratio of quinone to quinol was altered in mutants that have different expression levels of NemA, and physiological changes of quinones and glyoxals are associated such th...