Study Of Antimicrobial Peptides And Effect On Insulin Receptor Gene Of The Mouse Liver Cell Dealt With Glucose Tolerance Factor

Antimicrobial peptides are the immune response products of animals infected by the microbes. The antimicrobial pptides have many characters such as low molecular weight, thermostability, strong alkalinity and wide antibiogram. The ion channels on the cell membrane can be formed because of their distinctive mechanisms of killing microbes, and that cause the cell content to leak out and finally lead the bacteria to die. Antibiotic is the main way to deal with the diseases caused by the pathogens nowadays, but the resistances of microbes to the antibiotics are becoming the more and more serious problem that cannot be ignored. The resistance of microbes to antimicrobial peptides cann't be formed because of their special mechanisms of killing the microbes. So the antimicrobial peptides are the most prospective substitutes for the antibiotics. Because it is difficult to obtain antimicrobial peptides in large scales from animal tissues, the production of antimicrobial peptides by genetic engineering becomes the focus of people's interests.The subject in this study is the recombination , expression, purification and biological activity on antibacterial fusion peptide of, Magainin-Protegrin . Magainin is the skin secretion product of Batrachia under the eriviroment pressure, which can be used to accelerate the fusion rate of wound and resist the infection. It can inhibit the growth of bacteria and fungus with the lowest concentration; Protegrin is isolated from porcine leukocyte hi 1993 and is useful as antiviral agents.Firstly, the amino acid sequence of major bioactivity hi Magainin and Pro-tegrinin have been founded from the Genebank and then the related gene has been artifically synthesized. The enzyme sites are added to the two sides of the synthesized gene. The two peptides are connected with the three amino acid after the analysis of the software on the predication of the protein structure to avoid the interference hi the space. The PCR product was cut by two restriction enzymes, purified and cloned into the vector pGEX-4T-1. The engineering strains were incubated at 37 until OD600 ,0.6-0.8, then IPTG was added to reach the concentration of 0.75mmol/L to induce the fusion expression of the functional gene at 37 for 7 h. Tris-Gly-SDS-PAGE assay suggested the bacteria harboring the recombinant plasmid pGEX-4T-l- MP produce a fusion protein of 31KD. The purity of expressed protein after treatment by GST affinity chromatography reached 90% and at last the assay of inhibiting bacteria has been conducted.We also studied the effect on gene expression of the mouse liver cell which was dealt with the medicine of capsule of Fufang Gegen (the major component is the Glucose Tolerance Factor GTF). The main composition -chromium(3+) plays the important part in the increasement of insulin activity ,the enhencement of metabolism rate of sugar and fat , and the combination of insulin and the receptor of membrane. RNA has been extracted from the liver cell and the expression has been analyzed by the half-quantum after RT-PCR. The result displays the good effect of the medicine.