Sub-clining And Expression Of The Mae Gene Of The Hybrid Antibacterial Peptide MAE And Studies On Its Antibacterial And Anticancer Activities

MAE designed by our lab is a synthesized hybrid antibacterial peptide which possessed ideal antibacterial activity, it derives from the NH2-terminus of magainin and the NH2-terminus of melittin and these two regions are linked by a hinge region Ala-Gly-Pro.The mae gene(8lbp) has been synthesized based on the amino acid sequence of MAE and been cloned in the plasmid pTYB2 which is expressed in the E.coli ER2566.But the product of these syetem is not easily checked because of its scarcity and its low molecular weight. In this article the mae gene has been sub-cloned from pTYB2-mae to pMYB5 to solve this problem. Designing the upstream primer and the downstream primer, doubly digesting the PCR products and the plasmid pMYB5 with restriction enzymes XhoI and KpnJ , inserting with a single orientation, the gene mae has been successfully subcloned to pMYB5 by screening the transformants and sequencing. MBP-MAE would be purified from the induced expression product by flowing through the chitin affinity chromatography column with the lysate ER2566(pMYB5-MAE).MBP-MAE is easily checked even the scarcity of the expression because of its large molecular weight.MAE can be purified from the products of MBP-MAE cutted by CNBr through CM(carhoxymethyl) cellulose ion exchanger column according to its positive ion peptide property. MAE possesses antibacteria and anticancer activity.In this article the synthesized MAE was used to do the experiment on antibacteria and anticancer activity.The inhibition zone test of 16 clinical pathogenic bacteria show that MAE can strongly inhibit many Gram-positive and Gram-negative bacteria, it is possible that it will become a kind of broad-spectrum antibacterial agent.The lowest concentration may be 37.5 ug/mI.MAE can act on the nasopharyngeal carcinoma cells CNE-2 .Using the scanning electron microscope it was observed that the membrane structure of some areas was broken and injured and intracellular substances leaked out.The MTT test showed that the higher the concentration of MAE is, the stronger it acts on.