Acipensins - Novel Antimicrobial Peptides from Leukocytes of the Russian Sturgeon Acipenser gueldenstaedtii.

Antimicrobial peptides (AMPs) play an important role in the innate defense mechanisms in humans and animals. We have isolated and studied a set of antimicrobial peptides from leukocytes of the Russian sturgeon Acipenser gueldenstaedtii belonging to a subclass of chondrosteans, an ancient group of bony fish. Structural analy- sis of the isolated peptides, designated as acipensins (Ac), revealed in leukocytes of the Russian sturgeon six novel peptides with molecular masses of 5336.2 Da, 3803.0 Da, 5173.0 Da, 4777.5 Da, 5449.4 Da, and 2740.2 Da, designated as Ac1-Ac6, respectively. Complete primary structures of all the isolated peptides were determined, and the biologi- cal activities of three major components - Ac1, Ac2, and Ac6 - were examined. The peptides Ас1, Ас2, Ас3, Ас4, and Ac5 were found to be the N-terminal acetylated fragments 1-50, 1-35, 1-49, 1-44, and 1-51 of the histone Н2А, re- spectively, while Ас6 was shown to be the 62-85 fragment of the histone Н2А. The peptides Ac1 and Ac2 displayed potent antimicrobial activity towards Gram-negative and Gram-positive bacteria (Escherichia coli ML35p, Listeria monocytogenes EGD, MRSA ATCC 33591) and the fungus Candida albicans 820, while Ac6 proved effective only against Gram-negative bacteria. The efficacy of Ac 1 and Ac2 towards the fungus and MRSA was reduced upon an increase in the ionic strength of the solution. Ac1, Ac2, and Ac6, at concentrations close to their minimum inhibi- tory concentrations, enhanced the permeability of the E.coli ML35p outer membrane to the chromogenic marker, but they did not affect appreciably the permeability of the bacterial inner membrane in comparison with a potent pore-forming peptide, protegrin 1. Ac1, Ac2, and Ac6 revealed no hemolytic activity against human erythrocytes at concentrations of 1 to 40 μM and had no cytotoxic effect (1 to 20 μM) on K-562 and U-937 cells in vitro. Our findings suggest that histone-derived peptides serve as important anti-infective host defense molecules. k E y WORDS innate immunity; antimicrobial peptides; sturgeon leukocytes; histone H2A derivatives; acipensin. ABBREVIATIONS Ac - acipensin, AMP - antimicrobial peptides, CFU - colony forming units, EDTA - ethylenedi- aminetetraacetic acid, HNP - human neutrophil peptide (alpha-defensin), MIC - minimum inhibitory concen- tration, MRSA - methicillin resistant Staphylococcus aureus, ONPG - ortho-nitrophenyl β-D-galactopyranoside, PAGE - polyacrylamide gel electrophoresis, PBS - phosphate buffered saline, PCR - polymerase chain reac- tion, PG-1 - protegrin 1, SDS - sodium dodecyl sulfate, TFA - trifluoroacetic acid, TSB - trypticase soy broth; MALDI-TOF-MS - matrix-assisted laser desorption ionization time of flight mass spectrometry; Tris - tris (hydroxymethyl) aminomethane.