ABSTRACT
In hemolymph of insect species, compounds with remarkable properties for pharmaceutical industry are present. At the first line, there were found compounds of low molecular mass, less than 1 kDa. One of such compounds, ß-alanyl-tyrosine (252 Da), was isolated from larval hemolymph of some species of holometabolous insects (e.g. Neobellieria bullata). Its paralytic activity and antimicrobial properties were described until now. In this study, we present the effect of elongation of ß-alanyl-tyrosine by repeating of this motive on the biological and physical properties of prepared analogues. For assessment of antimicrobial properties of these new compounds strains of Gram-positive, Gram-negative bacteria and fungi were used, we also followed the haemolytic activity and toxic effect on human cell culture HepG2. On the base of ECD spectroscopy measurement, subsequent molecular modelling and known secondary structure of original ß-alanyl-tyrosine dipeptide, the secondary structures of repeating sequences of ß-AY were specified. The repeating structures of ß-alanyl-tyrosine show increase in antimicrobial activity; for Escherichia coli, Staphylococcus aureus and Pseudomonas aeruginosa, minimal inhibitory concentration was decreased from 30 to 15 mM for 2xß-AY, 0.4 mM for 4xß-AY and 0.25 mM for 6xß-AY.
Subject(s)
Dipeptides/chemistry , Dipeptides/pharmacology , Fungi/drug effects , Staphylococcus aureus/drug effects , Toxins, Biological/chemistry , Amino Acid Motifs , Anti-Infective Agents/chemistry , Anti-Infective Agents/pharmacology , Cell Proliferation/drug effects , Hep G2 Cells , Humans , Microbial Sensitivity Tests , Toxins, Biological/pharmacologyABSTRACT
We chose the larvae of fleshfly Sarcophaga bullata to map the peptide and protein immune response. The hemolymph of the third-instar larvae of S. bullata was used for isolation. The larvae were injected with bacterial suspension to induce an antimicrobial response. The hemolymph was separated into crude fractions, which were subdivided by RP-HPLC, gel electrophoresis, and free-flow electrophoresis. In several fractions, we determined significant antimicrobial activities against the pathogenic bacteria Escherichia coli, Staphylococcus aureus, or Pseudomonas aeruginosa. Among antimicrobially active compounds we identified dipeptide beta-alanyl-L-tyrosine, protein transferrin, and two variants of peptide sapecin. We also partially characterized two novel antimicrobially active polypeptides; odorant-binding protein 99b, and a peptide which remains unidentified.
