In vitro biosynthesis of autoinducer 2 of Streptococcus mutans UA1 59 and observation of the influence factors / 实用口腔医学杂志

ABSTRACT

Objective:To synthesize autoinducer-2 by the clone and prokaryotic expression of Streptococcus mutans(S.mutans)UAl59 luxS gene and to observe the influence factors.Methods:The expression vector pET21 a(+)-luxS of S.mutans UAl59 was transformed into Escheriehia coli BL2l(DE3).The S-ribosylhomocysteinase(Luxs)expression was induced by IPTG.The His tag fusion protein was isolated by Ni-chelating column and identified by Western blotting.Finally the protein was renatured by dialysis method.S-ribosylhomo-cysteine (SAH)was catalyzed by s-adenosylhomocysteine nucleosidas (Pfs)and LuxS,and then AI-2 was syntheszed.The AI-2 activi-ty was examined by luminescence of Vibrio harveyi BB1 70 when the concentration of LuxS protein or pH(4 -1 2)or the concentration of sodium fluoride was changed in reaction mixes of AI-2 synthesis.Results:Compared with the control group,with the increase of LuxS protein concentration,the relative activity of in vitro synthesized AI-2 increased gradually(P <0.001 ).When pH was between 6 -1 0, the relative activity of AI-2 were the highest,beyond the range of pH,the relative activity of AI-2 decreased(P <0.001 ).When a final concentration of sodium fluoride was more than 0.3%,the luminescence values decreased(P <0.05).Conclusion:LuxS fusion protein can promote the production of AI-2.Optimum pH for AI-2 biosynthesis in vitro must be between 6-1 0.Biosynthesis of AI-2 is inhibited by sodium fluoride with final concentration of more than 0.3%.