The Preparation And Structure-Activity Relationship Of Antimicrobial Polypeptides

Since the discovery of penicillin in 1928,antibiotics have played an important role in the field of medical care.However,due to the abuse of antibiotics,the drug resistance of micribials is growing steadily,which poses serious threat to human health.According to the prediction of the World Health Organization,the number of deaths caused by drug-resistant microbial infection will exceed that of cancer by 2050.Therefore,there is an urgent need to develop new antibiotics with broad-spectrum antibacterial properties and not easy to cause microbial drug resistance.In the past three decades,antimicrobial peptides have been widely studied as potential substitutes for antibiotics because of their broad spectrum and effective antibacterial activity.However,the clinical application of antimicrobial peptides is greatly limited,due to the high production cost,low bioavailability,instability to protease,toxicity of systemic administration.In recent years,cationic polymers that mimic the structural of antimicrobial peptides(such as positive charge and hydrophobic structure)have shown great potential in the treatment of multidrug resistant microbial infections because of their advantages of easy synthesis and modification,broad-spectrum antibacterial activity,low cytotoxicity and stability to protease degradation.In contrast to traditional small molecule antibiotics,cationic polymers can destroy the integrity of the microbial cell membrane and causes effective microbial death,and is not likely to develop microbial drug resistance.However,the strong toxicity to mammalian cells greatly limited the application of cationic polymers.Therefore,it is urgent to develop novel antimicrobial materials with good antimicrobial activity and negligible toxicity to host cells.In this paper,a series of polypeptides with different structures were obtained by ring opening polymerization and post polymerization modification of N-carboxylic anhydride(NCA)monomer.The effects of polypeptides’structure on their antimicrobial activity and biocompatibility were systematically studied.The following research results were obtained:1.Low-Molecular-Weight Polylysines with Excellent Antibacterial Properties and Low Hemolysis.Firstly,the ring opening polymerization of lysine NCA was initiated by different hydrophobic primary amines to obtain side chain benzyloxycarbonyl protected polylysine.Then,a series of low molecular weight cationic polylysine(C_x-PLL_n)with different hydrophobic end groups(C_x)and degree of polymerization(PLL_n)were obtained by deprotection reaction.In vitro antibacterial experiments showed that the synthesized polypeptides shown good antibacterial activity.Mechanism studies shown that this kind of homolysine can effectively destroy the integrity of bacterial cell membrane and lead to bacterial death.Owing to this membrane-disrupting property,this kind of polylysine showed rapid bacterial killing kinetics and was not likely to develop resistance after repeat treatment(up to 13 generations).Moreover,C_x-PLL_ndemonstrated a significant therapeutic effect on an MRSA infection mouse model,which further proved that this synthetic polymer could be used as an effective weapon against bacterial infections.2.Photosensitizer-polypeptide conjugate for effective elimination of Candida albicans biofilm.Persistent fungal infection caused by fungal biofilm is a serious threat to human health.Therefore,in the second part of this work,a photosensitizer-polypeptide conjugate(PPa-cP)comprising a photosensitizer,pyropheophorbide a(PPa),and a cationic polypeptide(cP)was readily synthesized for effective antifungal and antibiofilm treatment.Compared with the free photosensitizer PPa,the cationic conjugate can effectively bind with the negatively charged Candida albicans(C.albicans)membrane through electrostatic action,thus enhancing the enrichment on the surface of fungi,and showing effective antifungal effect on C.albicans and fluconazole resistant C.albicans under light irradiation.In addition,the electrostatic interaction between the PPa-cP conjugate and fungi enhanced the penetration in C.albicans biofilm.Therefore,this kind of PPa-cP conjugate can effectively eliminate the C.albicans biofilm under light irradiation.More importantly,PPa-cP demonstrated significantly enhanced therapeutic effects in a fluconazole-resistant C.albicans-infected rat model with minimal side effects.Therefore,this part of work provides an effective strategy to combat biofilm infections associated with biomedical equipment.3.Photosensitizer-Polypeptides Conjugate with Synergistic Antibacterial Efficacy.Based on the work of the previous chapter,in order to further evaluated the effects of side chain cationic groups of conjugates on their antibacterial properties and hemolysis,and improve their biocompatibility and application potential in vivo,a series of quaternary ammonium salt functionalized photosensitizer polypeptide conjugate(PPa-cP)were obtained through NCA ring opening polymerization and post modification.Compared with free PPa,the cationicPPa-cP showed enhanced binding ability with the negative surface of S.aureus through electrostatic interaction,exhibiting effective antibacterial activity against both S.aureus and MRSA in vitro under light irradiation.Of the synthesized PPa-cP,PPa-cP5 with the degree of polymerization of 37 and modified with the 1-Methylimidazole side group exhibited the best antibacterial activity with a minimum inhibitory concentration(MIC)value of2μM without light irradiation and 0.25μM with light irradiation,respectively.Moreover,the PPa-cP5 showed good hemocompatibility.The above results elucidate that the positively charged PPa-cP5 could significantly increase the efficiency of PDT and effectively eradicate S.aureus biofilm due to its potent penetration ability into S.aureus biofilms.Overall,the present study establishes an efficient strategy to resist S.aureus and S.aureus biofilm infections.