Abstract
Beta-defensins are important part of innate immunity of fish, which are the first defense line against invading pathogens. In this study, the β-defensin (Lhβ-defensin) gene was cloned from spleen tissue of soiny mullet (Liza haematocheila). Lhβ-defensin cDNA was 747 bp in length, encoding 63 amino acids. Sequence alignment revealed that Lhβ-defensin contained six conserved cysteine residues and shared 97.5% sequence identities with grouper (Epinephelus coioides) β-defensin. Realtime PCR revealed that Lhβ-defensin was highest expressed in the immune related organs, such as spleen, kidney and gut of healthy fish. Following Streptococcus dysgalactiae infection, Lhβ-defensin was up-regulated in immune related organs, e.g. 17.6-fold in spleen and 10.87-fold in gut at 24 h post infection (hpi). Lhβ-defensin possessed a monomeric structure of a three-stranded anti-parallel β-sheet and an α-helix stabilized by three disulfide bonds formed by Cys30-Cys58, Cys36-Cys52, and Cys40-Cys59. In addition to the experimental work, computer simulation was also carried out to determine the possible conformation of β-defensin and its interaction with palmitoyloleoylphosphatidylglycerol (POPG), a model of bacteria membrane. The Lhβ-defensin was found to form dimeric structure stabilized by the van der Waals contacts of Leu35 and Cys37 in two anti-parallel β1-strands and the cation-π interaction between Tyr32 and Arg54 respectively in the two β1-strands. The most important interactions between β-defensin and membrane are the electrostatic interactions between Arg residues in β-defensin and head group of POPG bilayer as well as hydrogen bond interactions between them. Our results were useful for further understanding the potential mechanism of antimicrobial property of fish β-defensins.
Highlights
Antimicrobial peptides (AMPs) are important components of the innate immunity acting as the first defense line against invading pathogens
The defensins are cysteine-rich cationic antimicrobial peptides which are divided into three distinct groups, α, β- and θ-defensins, based on the different disulfide bond [1,2]. α-defensins were isolated from mammalian neutrophils and macrophages which are stabilized by a mode of disulfide bonds of C1-C6, C2-C4 and C3-C5. θ-defensins are the macrocyclic product of a head-to-tail of two truncated α-defensins which are only found in leukocytes and bone marrows of Old World monkeys [2, 3]. β-defensin are the largest group of defensins family and are the most characteristic defensins. β-defensins are comprised of 35 to 50 amino acids with a core of three anti-parallel β-strands stabilized by a mode of disulfide bond of C1-C5, C2-C4 and C3-C6 [4]
Β-defensins had been characterized in a number of fish species, such as zebrafish (Danio rerio), fugu (Takifugu rubripes), tetraodon (Tetraodon nigroviridis) [5], blunt snout bream (Megalobrama amblycephala) [6] and Nile tilapia (Orechromis niloticus) [7]
Summary
Antimicrobial peptides (AMPs) are important components of the innate immunity acting as the first defense line against invading pathogens. The interaction of Lhβ-defensin dimer and bacterial membrane (POPG lipid membrane) was investigated base on the MD simulation. Interaction between Lhβ-defensin dimer and POPG bilayer
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