Transcriptional Activation of the Aldehyde Reductase YqhD by YqhC and Its Implication in Glyoxal Metabolism of Escherichia coli K-12

Abstract

The reactive ␣-oxoaldehydes such as glyoxal (GO) and methylglyoxal (MG) are generated in vivo from sugars through oxidative 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 (GO r ) mutants from Escherichia coli strain MG1655 on LB plates containing 10 mM GO. By tagging the mutations with the transposon TnphoA-132 and determining their cotransductional linkages, we were able to identify a locus to which most of the GO r mutations were mapped. DNA sequencing of the locus revealed that it contains the yqhC gene, which is predicted to encode an AraC-type transcriptional regulator of unknown function. The GO r mutations we identified result in missense changes in yqhC and were concentrated in the predicted regulatory domain of the protein, thereby constitutively activating the product of the adjacent gene yqhD. The transcriptional activation of yqhD by wild-type YqhC and its mutant forms was established by an assay with a ␤-galactosidase reporter fusion, as well as with real-time quantitative reverse transcription-PCR. We demonstrated that YqhC binds to the promoter region of yqhD and that this binding is abolished by a mutation in the potential target site, which is similar to the consensus sequence of its homolog SoxS. YqhD facilitates the removal of GO through its NADPH-dependent enzymatic reduction activity by converting it to ethadiol via glycolaldehyde, as detected by nuclear magnetic resonance, as well as by spectroscopic measurements. Therefore, we propose that YqhC is a transcriptional activator of YqhD, which acts as an aldehyde reductase with specificity for certain aldehydes, including GO.

YqhD: a broad-substrate range aldehyde reductase with various applications in production of biorenewable fuels and chemicals

“…Similar results were observed for glyoxal and methylglyoxal: deletion of yqhD increased sensitivity and increased YqhD expression increased tolerance (Lee et al 2010). Alternatively, sensitivity to Cd 2+ or diamide, which do not generate reactive oxygen species, was not altered by changes in yqhD expression (Perez et al 2008).…”

“…Contrastingly, when Riehle et al evolved three parallel E. coli lines for thermotolerance, yqhD transcript abundance was increased approximately 4-fold in two of these lines relative to the parental E. coli B-derived strain (Riehle et al 2003;Riehle et al 2005). Finally, glyoxaltolerant mutants of MG1655 were found to have increased expression of yqhD (Lee et al 2010). …”

“…YqhD has NADPH-dependent reductase activity towards many of these toxic aldehydes, such as acrolein, butyraldehyde and glyoxal (Table 1) (Perez et al 2008;Lee et al 2010). This activity plays a defensive role against reactive oxygen species-generating agents, such as hydrogen peroxide and potassium tellurite: deletion of yqhD increased growth sensitivity and increased expression of YqhD decreased growth sensitivity (Perez et al 2008).…”

“…This perturbed expression of yqhD was shown to be critical for furfural and glyoxal tolerance (Lee et al 2010;Miller et al 2009b), but the contribution of increased yqhD expression to thermotolerance has not been determined.…”

“…Reverse engineering efforts revealed that both the furfural-tolerant and glyoxal-tolerant strains contained a critical mutation within yqhC, a putative regulator that is adjacent to yqhD, though controlled by a separate promoter Lee et al 2010). The furfuraltolerant strain contained an insertion mutation, rendering YqhC unable to activate yqhD ).…”