ABSTRACT
ABSTRACT This study aimed to describe a Bacillus subtilis expression system based on genetically modified B. subtilis. Abaecin, an antimicrobial peptide obtained from Apis mellifera, can enhance the effect of pore-forming peptides from other species on the inhibition of bacterial growth. For the exogenous expression, the abaecin gene was fused with a tobacco etch virus protease cleavage site, a promoter Pglv, and a mature beta-glucanase signal peptide. Also, a B. subtilis expression system was constructed. The recombinant abaecin gene was expressed and purified as a recombinant protein in the culture supernatant. The purified abaecin did not inhibit the growth of Escherichia coli strain K88. Cecropin A and hymenoptaecin exhibited potent bactericidal activities at concentrations of 1 and 1.5 µM. Combinatorial assays revealed that cecropin A and hymenoptaecin had sublethal concentrations of 0.3 and 0.5 µM. This potentiating functional interaction represents a promising therapeutic strategy. It provides an opportunity to address the rising threat of multidrug-resistant pathogens that are recalcitrant to conventional antibiotics.
Subject(s)
Antimicrobial Cationic Peptides/genetics , Antimicrobial Cationic Peptides/metabolism , Bacillus subtilis/genetics , Genetic Vectors/genetics , Insect Proteins/genetics , Insect Proteins/metabolism , Anti-Bacterial Agents/isolation & purification , Anti-Bacterial Agents/metabolism , Anti-Bacterial Agents/pharmacology , Antimicrobial Cationic Peptides/isolation & purification , Antimicrobial Cationic Peptides/pharmacology , Bacillus subtilis/metabolism , Escherichia coli/drug effects , Escherichia coli/growth & development , Gene Expression , Genetic Vectors/metabolism , Insect Proteins/isolation & purification , Insect Proteins/pharmacology , Protein Engineering , Protein Transport , Recombinant Proteins/genetics , Recombinant Proteins/isolation & purification , Recombinant Proteins/metabolism , Recombinant Proteins/pharmacologyABSTRACT
In this study, anticoagulant activity was detected in salivary gland homogenates (SGHs) of Thyrsopelma guianense (Diptera: Simuliidae). The SGH yielded 1.07 ìg ± 0.03 (n = 15) of total soluble protein per pair of glands. In addition, following SDS-PAGE (12.5 percent gel) and silver nitrate staining, 12 polypeptides with molecular weights ranging from 14-69 kDa were detected in all physiological ages analyzed (12 h, 24 h, 48 h and 72 h following emergence). Coagulation bioassays showed that the SGHs had activities that interacted at all levels of coagulation (the intrinsic, extrinsic and common pathways), by extending the plasma recalcification time, prothrombin time, thrombin time. This is the first report on the activity of salivary gland proteins from the main vector of onchocerciasis in Brazil. We also suggest detailed studies on the morphology and function of the salivary glands in order to understand the role of these proteins in host/vector interactions.