Effects Of Terminal Amino Acids Of Alpha Helical Peptides On Their Structure And Biological Activity

In recent years,bacteria resistance becomes more and more serious because of the abuse of antibiotics,while the incidence and mortality of cancer are very high,the traditional antibacterial and anticancer drugs had been unable to meet human needs.Due to the use of common antibiotics and antitumor drugs,drug resistance of bacteria and tumor cells is serious.To solve the current predicament,people need to develop a new drug which has high efficiency and low toxicity,broad scope and uneasy to produce drug resistance.Antimicrobial peptides are widely distributed in eukaryotic organisms,including mammals,amphibians,insects,plants and animal in prokaryotes,They are a class of peptides generated in the exogenous pathogen invasion,composed of 30⁶0 amino acids,with broad spectrum,high efficiency and specificity against exogenous pathogens.The amphipathic ?-helical antimicrobial peptides are widely distributed,has simple structure and easy to synthesis,so they are usually used as model molecules for study on the relationship between structure and function of peptides.Previous studies had found that G?IIKK?₃I-NH₂ had good antibacterial and antitumor activity,but less toxicity to normal cells.In addition,remove the N-terminal or C-terminal amino acid had a great influence on the biological activity of the antimicrobial peptide.This dissertation used G?IIKK?₃I-NH₂ as template,based on the nature of the different amino acids,we designed and synthesized three groups of polypeptides by modifying the N terminal or C terminal amino acid residues: 1)replace the N terminal amino acid: A?IIKK?₃I-NH₂,E?IIKK?₃I-NH₂,K?IIKK?₃I-NH₂,V?IIKK?₃I-NH₂;2)replace the C terminal amino acid: G?IIKK?₃A-NH₂,G?IIKK?₃E-NH₂,G?IIKK?₃KNH₂,G?IIKK?₃V-NH₂;3)add or remove terminal amino acid: G?IIKK?₃II-NH₂,AA?IIKK?₃I-NH₂,GA?IIKK?₃I-NH₂,?IIKK?₃II-NH₂.In the three groups of peptides,we replaced the terminal amino acid glycine?G?by alanine?A?and valine?V?,whose hydrophobic side chain residues change little,the positively charged lysine?K?and negatively charged glutamic acid?E?,we also added or removed terminal amino acids.We examined the modified effects of the peptides on their structure and biological activity.We used CD to detect the change of secondary structure under different conditions,and we found that these peptides were random coil in the hydrophilic environment while transformed to ?-helical structure in the hydrophobic environment.This amphipathic structure made it easier to insert the bacteria and tumor cell membrane,causing membrane repute,then exerted antimicrobial and antitumor activity.Through comparison of various properties of peptides,including the retention time in HPLC,antibacterial activity,anti-tumor activity,toxicity to normal cells,the immune response of lymphocytes,the destructive effect to tumor cells membrane and the membrane insertion ability,we found that the replacement of N terminal amino acid had little effect on the activity of the peptide,but the biological activity of the peptides had obvious differences after replacing C terminal amino acid,and presented a consistent rule: G?IIKK?₃I-NH₂>G?IIKK?₃V-NH₂>G?IIKK?₃A-NH₂>G?IIKK?₃ENH₂>G?IIKK?₃K-NH₂.In addition,when the C-terminal had 2 hydrophobic amino acids isoleucine?I?,the biological activity of peptides increased significantly,but when the N-terminal had 1 or 2 hydrophobic amino acids alanine?A?,biological properties had no significant change,indicating that the C terminal amino acid plays a more important role in G?IIKK?₃I-NH₂.This dissertation also discussed the G?IIKK?₃I-NH₂ whether can lead to the bacteria?ESBL E.coli?resistance,after 30-40 repeated dosing,G?IIKK?₃I-NH₂ didn't cause bacterial resistance.Through the detection of resistance genes,we also found that the expression of drug resistance gene treated by G?IIKK?₃I-NH₂ was lower than that with antibiotics.