Rational Design,Activity Evaluation And Mechanism Study Of Small Peptides Against Acinetobacter Baumannii

Acinetobacter baumannii(AB)is a type of common gram-negative bacilli,which is distributed in hospitals worldwide and survive for long time,and is an opportunistic nosocomial pathogen which is responsible for a vast array of infections including respiratory infections,bacteremia,secondary meningitis,urinary tract infections,surgical site infections.This human pathogen has emerged as one of the most troublesome pathogens threatening the current antibiotic era for health care institutions globally which is propelled by the use of broad-spectrum antimicrobial agents and its remarkable ability to up regulate or acquire resistance determinants.The drug resistance of Acinetobacter baumannii possess various mechanisms,manifesting as multidrug resistance or even pan-drug resistance with an obvious increasing trend of tolerance.In view of the serious state of antibiotic against acinetobacter baumannii,it is becoming a current focus on developing new anti-infective agents.Antimicrobial peptides(AMPs)are an integral part of the innate immune system that protects a host from the invasion of pathogenic bacteria.AMP has a variety of mechanisms involved to antimicrobial effects,which to be currently reported is disrupting the membrane,inhibiting cell-wall synthesis,altering cytoplasmic membrane,inhibiting enzymatic activity,inhibiting DNA,RNA and protein synthesis,and binding to DNA.In view of the antibacterial mechanism of antibacterial peptide is different from the antibacterial mechanism of antibiotic,AMPs have many advantages including broad spectrum,rapid sterilization and low drug resistance,especially good antibacterial ability for drug-resistant bacteria,AMPs are expected to become an ideal substitute for antibiotics.By linking up the research with rational design-activity evaluation-mechanism research as a main thread,non-helical peptides and helical peptides against acinetobacter baumannii are studied.The main achievements of the article are as follows:1.Optimal design methods for AMP structure are established and four non-helical peptides and two helical peptides are designed and obtained.2.Novel AMPs have antibacterial effect upon gram-positive and gram-negative bacteria,especially resistant Acinetobacter baumannii.The minimum inhibitory concentrations(MICs)of non-helical peptide A4 and helical peptide B2 against resistant strains of Acinetobacter baumannii are both 8?g/m L,and are 8?g/m L and 4?g/mL against multi-drug resistant strains of Acinetobacter baumannii respectively,which show comparative antibacterial activity with the imipenem in vitro.3.Novel AMPs can delay growth of multi-drug resistant strains of Acinetobacter baumannii.When the number of bacteria is more than 10~3 CFU/mL on the culture medium,the germicidal speeds of non-helical peptide A2,A4 and helical peptide B2greater than imipenem.4.Concentration and charge of salt ions can affect the biological activity of the novel antibacterial peptide.Peptides A3 and A4 possess good plasma stability,and the modification of phenylethyamine at the C-fragment of the polypeptide chain can improve the stability of newly designed non-helical peptides in plasma.5.Non-helical peptides A2,A3,A4 and A5 have low hemolytic toxicity and high antibacterial activity,while helical peptide B2 have high hemolytic toxicity and need to be optimized further.Peptides A4,A6 and B2 have strong anti-tumor activity in vitro,and more than 80%Hep G2 cells are killed when the concentrations of three peptides are 256?g/m L.6.Non-helical peptideA2,A4 and helical peptide B2 combined with imipenem respectively have synergy effect on multi-drug resistant strains of Acinetobacter baumannii and the synergy effect of A2 and A4 is better than B2.7.Through a series of experiments including LSCM,Flow cytometer,SEM,the liposome experiment and the gel retardation experiment,it is determined primarily that the targets of the antibacterial action of the peptides are cell membrane and DNA for newly designed non-helical peptides,are mainly DNA for newly designed helical peptides.Of course,other potential antibacterial targets may exist and it is remained to be studied further.