Study On Response Of E. Coli To Mercury And Selenium Exposure By Analytical Approach To The Metalloproteomics

The response of organisms to selenium or/and mercury exposure was investigatrd usinghigh-throughout and comprehensive proteomics and metalloproteomics methods with E. coli.as a research model. The growth curve was taken out by absorbance measurements to studythe effect of mercury and/or selenium on E. coli. The total proteins were separated usingisoelectrofocusing/sodium dodecyl sulfate-polyacrylamide gel electrophoresis(IEF/SDS-PAGE), a method with high resolution. Parallel experiments were designed, onegroup of gels were dyed with Coomassie brilliant blue R-250, following by PDQuest softwareanalysis; the other group of gels were fixed and then were dried immediately. Dried gels weresubject to elemental mapping analyzed employing advanced nuclear analysis technique ofsynchrotron radiation X-ray fluorescence (RSXRF). The related important proteins werecharacterized with electrospray ionization tandem mass spectrometry (ESI-MS/MS) in orderto detect the proteins containing Se and/or Hg and the responded proteins. The responsemechanism of E. coli to mercury exposure and the antagonistic mechanism of selenium andmercury were studied on this base. The results were listed as following:â‘ The Lag phase was prolonged to22h when exposed to mercury only, indicating thetoxicity of mercury. The lag phase was shorten to18h when added selenium tomercury-containing medium, implying the antagonism of selenium to mercury toxicity.â‘¡Separated dyed gels were analyzed by PDQuest,5proteins were detected withsignificant differential expression, and one selenium-and mercury-binding protein wasdetected with elemental analysis. The above mentioned six proteins were characterized, andthe significant different proteins were Outer membrane protein W (OmpW), Transcriptiontermination factor rho, Cysteine synthase (Cys), Transaldolase A (Tal) and Alkylhydroperoxide reductase subunit C (AhpC) respectively, and the se-and Hg-containingprotein was pyruvate kinase (PK). Of the six proteins, OmpW was of down regulation in E.coli cultured in Hg-containing medium, and the others were of up regulation when comparedwith control. OmpW expression was higher, and Tal expression was lower when added selenium to Hg-exposed culture.â‘¢Combined with the biological function of these proteins, the response mechanism of E.coli to mercury exposure was clear: by up-regulating the expression of rho, the biosynthesiswas controlled and the penetrability of Hg was reduced. Cells were protected from injury bymercury; by up-regulating the expression of AhpC, superfluous free radical induced bymercury was deoxidized, reducing the oxidized damnify; by being bonded with mercury,some enzymes (such as PK) with hydroxyl were inactive, inducing metabolic disturbance. Inorder to keep the normal metabolic activity, other enzyme with the same function wasexpressed up-regulated. The antagonistic mechanism of selenium to mercury toxicity ismanifested in two aspects: species containing selenium combined with mercury to acompound which was nontoxic, improving the penetrability induced by mercury in some way,Down-regulating the expression of rho and up-regulating the expression of OmpW relatively;combination of selenium species to mercury reduced that of mercury to hydroxyl in enzymes(PK), and led to down-regulated expression of other enzyme (Tal) with the same function.