Construction Of Multi-drug Resistance Staphylococcus.aureus And Chinese Herbal Medcine Inhibitor Of Efflux Pump

Multi-drug Efflux Pumps is an important mechanism in multi-drug resistance Staphylococcus.aureus. To characterize the norA gene and bmr gene which conferred relatively high resistance to hydrophilic quinolones in S.aureus, We sequenced the genes and analyzed the deduced polypeptide. Then, the bmr gene was over expressed in Escherichia coli cells. We constructed shuttle plasmid pUTnorA and pUTbmr, which is composed of the vector pUB-110 and pMD-18T containing a strong promoter and bmr or norA gene. Quinolone-susceptible S.aureus was transformed with the resulting plasmid, pUTnorA and pUTbmr. MIC of ravious hydrophilic quinolones for the transformed S.aureus was 16 to 64 times greater to susceptible S.aureus. In conclusion, we have constructed multi-drug resistance S.aureus, in which was known about the genetics and mechanisms of quinolone resistance. It's used to select Chinese herbal medicine inhibitor of efflux pump. The nucleotide sequences of norA gene(1 317bp) and bmr gene(1 256bp) were determined. The norA sequence contained only one open reading fame (nucleotides 153-1 317bp) long enough to encode a polypeptide of 388 amino acids. Putative promoter sequences were found nucleotides (TACAAT and TTGACT), which well match the consensus sequences (TATAAT and TTGACA) for the -10 and -35 regions of E.coli promoters. Rather, the T→A norA promoter region mutation probably play a role in efflux-mediated multidrug resistance. The amino acid composition of the deduced NorA and Bmr polypeptides were shown they are rich in hydrophobic such as Ala, Ile, Leu, Phe, Trp and Val. NorA has 187 (48%) hydrophobic amino acids of the 388; Bmr has 196 (50%) hydrophobic amino acids of the 389.