超强静磁场对染料脱色菌Escheriehia coli的acpD基因的影响

将大肠杆菌E.coli ATCC8739置于12.0 T超强静磁场(ultra-strong static magnetic field,SMF)中进行处理,获得了磁场处理0.5、1、2、4和8 h的菌株E.coli-SMFn(n=0.5、1、2、4、8)。在37℃、pH 7、静置的条件下,菌株对偶氮染料AR14(I.C.Acid Red 14)的脱色结果指出,当反应进行到4、6和8 h时,E.coli-SMF8的脱色效率分别为18%、55%和96%,远高于E.coli ATCC8739的3%、19%和50%,表明SMF作用显著地增强了菌株的脱色效率。基因组DNA提取、PCR扩增、分子克隆以及基因测序的实验结果进一步表明,全部6例E.coli ATCC8739菌株的偶氮还原酶基因(acpD)序列均与GenBank中报道的完全一致;而E.coli-SMF8菌株的acpD-SMF8核酸序列中缺失了第99位的胞嘧啶。该缺失突变不仅使acpD-SMF8的核酸序列与acpD的存在显著不同,同时得到了具新活性中心的偶氮还原酶AzoR-SMF8。这个新的活性中心具有更强的黄素(FMN)结合能力,因此使该酶与偶氮染料的亲和力大大增加,促进了脱色效率的提高。

Mutagenesis effect of ultra-strong static magnetic field on acpD gene of Escheriehia coli

Magnetic treated strains E.coli-SMFn(n=0.5,1,2,4,8) were obtained when E.coli ATCC8739 was exposed to 12.0 T ultra-strong static magnetic field(SMF) for 0.5,1,2,4 and 8 h,respectively.All of them were used for the decolorization of Acid Red 14 under the static condition(37 ℃,pH 7).The results indicated that the decolorization rates of E.coli-SMF8 at 4,6 and 8 h were 18%,55%,and 96%,respectively,apparently much higher than those of E.coli ATCC8739(3%,19% and 50%).This suggested that the effect of SMF could significantly enhance the decolorization ability of E.coli.Then by means of genomic DNA extraction,PCR amplification,molecular cloning and gene sequencing,it was found that the nucleotide sequence of acpD belonged to the original strain was completely consistent with that reported in genbank,while the nucleotide sequence of acpD-SMF8 presented a cytosine deletion at the 99th point.The deletion mutation not only led to the change of nucleotide sequence,but also created a novel azoreductase(AzoR-SMF8).The active center of this novel enzyme possessed much stronger ability of binding FMN,which could multiply the affinity between azoreductase and azo dye,and enhance the decolorization efficiency.