ABSTRACT
Two families of structurally related C-terminally alpha-amidated antimicrobial peptides have been identified in norepinephrine-stimulated skin secretions of the midwife toad Alytes obstetricans (Alytidae). The alyteserin-1 peptides (Gly-Leu-Lys-(Asp/Glu)-Ile-Phe-Lys-Ala-Gly-Leu-Gly-Ser-Leu-Val-Lys-(Gly/Asn)-Ile-Ala-Ala-His-Val-Ala-(Asn/Ser).NH(2)) show limited structural similarity to the ascaphins from the skins of frogs of the family Leiopelmatidae. Alyteserin-2a (Ile-Leu-Gly-Lys-Leu-Leu-Ser-Thr-Ala-Ala-Gly-Leu-Leu-Ser-Asn-Leu.NH(2)) and alyteserin-2b and -2c (Ile-Leu-Gly-Ala-Ile-Leu-Pro-Leu-Val-Ser-Gly-Leu-Leu-Ser-(Asn/Ser)-Lys-Leu x NH(2)) show limited sequence identity with bombinin H6, present in the skins of frogs of the family Bombinatoridae. The alyteserin-1 peptides show selective growth inhibitory activity against the Gram-negative bacteria Escherichia coli (MIC=25 microM) whereas alyteserin-2a is more potent against the Gram-positive bacteria Staphylococcus aureus (MIC=50 microM). The hemolytic activity against human erythrocytes of all peptides tested is relatively weak (LC(50)>100 microM). The data demonstrate that the frogs belonging to the family Alytidae are among those producing dermal antimicrobial peptides that may represent a component of the animal's system of innate immunity.
Subject(s)
Anti-Bacterial Agents/pharmacology , Antimicrobial Cationic Peptides/pharmacology , Anura , Erythrocytes/drug effects , Escherichia coli/drug effects , Peptide Fragments/pharmacology , Skin/metabolism , Staphylococcus aureus/drug effects , Animals , Anti-Bacterial Agents/chemistry , Anti-Bacterial Agents/isolation & purification , Antimicrobial Cationic Peptides/chemistry , Antimicrobial Cationic Peptides/isolation & purification , Chromatography, High Pressure Liquid , Humans , Microbial Sensitivity Tests , Peptide Fragments/chemistry , Peptide Fragments/isolation & purification , Skin/chemistry , Time FactorsABSTRACT
Antimicrobial peptides in the skin secretions of anurans constitute a component of the innate immunity that protects the organism against invading pathogens. Four peptides with antimicrobial activity were isolated in high yield from norepinephrine-stimulated skin secretions of the Northern red-legged frog Rana aurora aurora and their primary structures determined. Ranatuerin-2AUa (GILSSFKGVAKGVAKNLAGKLLDELKCKITGC) showed potent growth-inhibitory activity against a range of Gram-positive and Gram-negative bacteria (minimum inhibitory concentrations < 20 microM) but low hemolytic activity against human erythrocytes (50% hemolysis at 290 microM). Brevinin-1AUa (FLPILAGLAAKLVPKVFCSITKKC) and brevinin-1AUb (FLPILAGLAANILPKVFCSITKKC) also showed potent antimicrobial activity but were strongly hemolytic (HC50 < 10 microM). Temporin-1AUa (FLPIIGQLLSGLL.NH2) atypically lacked a basic amino acid residue and showed very weak antimicrobial and hemolytic activity. Its biological function remains to be established. The primary structures of the antimicrobial peptides are consistent with a close phylogenetic relationship between R. aurora, Rana boylii and Rana luteiventris.
