Antibacterial activity of six novel peptides from Tityus discrepans scorpion venom. A fluorescent probe study of microbial membrane Na+ permeability changes

Six novel peptides (named bactridines) were isolated from Tityus discrepans scorpion venom. From mass spectrometry molecular masses were 6916, 7362, 7226, 7011, 7101 and 7173 Da (bactridines 1-6). Bactridines 1 and 2 were sequenced by Edman degradation. The sequences and in silico analysis, indicated that they are positively charged polypeptides comprised of 61 and 64 amino acids (AA), respectively, bactridine 1 and bactridine 2 containing 4 disulfide bridges. Bactridine 1 was only toxic to cockroaches and crabs, and bactridine 2-6 were only toxic to mice. Bactridine 1 has a 78% sequence identity with ardiscretin. Ardisctretin is an insect specific sodium toxin which also produces a small depolarization and induces repetitive firing in squid axons resembling those of DDT [1,10(pchlorobenzyl) 2-trichloretane] in its ability to slow down action potential, to induce repetitive firing. Measured as the minimal inhibitory concentration, bactridines had high antibacterial activity against a wide range of Gram positive and Gram negative bacteria. Complete bacterial growth inhibition occurred at concentrations from 20 to 80 mu M depending on the bacteria and peptide tested. Effects on membrane Na+ permeability induced by bactridines were observed on Yersinia enterocolitica loaded with 1 mu M CoroNa (TM) Red. CoroNa (TM) Red fluorescence leakage from bacteria was observed after exposure to 0.3 mu M of any bactridine tested, indicating that they modified Na+ membrane permeability. This effect was blocked by 10 mu M amiloride and by 25 mu M mibefradil drugs that affect Na+ and Ca2+ channels respectively. We found no evidence of changes of K+ or Ca2+-concentrations neither inside nor outside the bacteria in experiments using the fluorescent dyes Fluo 4AM (10 mu M) and PBFI (20 mu M). (C) 2009 Elsevier Ltd. All rights reserved.