SpALF4: A newly identified anti-lipopolysaccharide factor from the mud crab Scylla paramamosain with broad spectrum antimicrobial activity

• Published in: Fish & shellfish immunology Vol. 36; no. 1; pp. 172 - 180
• Main Authors: Zhu, Lei, Lan, Jiang-Feng, Huang, Yan-Qing, Zhang, Chao, Zhou, Jun-Fang, Fang, Wen-Hong, Yao, Xiao-Juan, Wang, Hao, Li, Xin-Cang
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 01.01.2014
• Subjects: Aeromonas hydrophila; Amino Acid Sequence; Animals; Antimicrobial activity; Antimicrobial Cationic Peptides - chemistry; Antimicrobial Cationic Peptides - genetics; Antimicrobial Cationic Peptides - immunology; Bacillus megaterium; Base Sequence; Binding activity; Brachyura - genetics; Brachyura - immunology; Candida albicans; Cloning, Molecular; Decapoda; Expression pattern; Gram-Negative Bacteria - immunology; Gram-Positive Bacteria - immunology; Hydrophobic and Hydrophilic Interactions; Isoelectric Point; Microbial Sensitivity Tests; Models, Molecular; Molecular Sequence Data; Nimaviridae; Phylogeny; Pseudomonas putida; Recombinant Proteins - genetics; Recombinant Proteins - immunology; Reverse Transcriptase Polymerase Chain Reaction; RNA - chemistry; RNA - genetics; Scylla paramamosain; Sequence Alignment; Sequence Analysis, DNA; SpALF4; Staphylococcus aureus; Staphylococcus aureus - immunology; Vibrio alginolyticus; Vibrio anguillarum; Vibrio harveyi; Antimicrobial activity; Expression pattern; SpALF4; Binding activity
• ISSN: 1050-4648; 1095-9947
• DOI: 10.1016/j.fsi.2013.10.023

Anti-lipopolysaccharide factors (ALFs) are antimicrobial peptides with binding and neutralizing activities to lipopolysaccharide (LPS) in crustaceans. This study identified and characterized a novel ALF homolog (SpALF4) from the mud crab Scylla paramamosain. The complete cDNA of SpALF4 had 756 bp with a 381 bp open reading frame encoding a protein with 126 aa. The deduced protein contained a signal peptide and a LPS-binding domain. SpALF4 shared the highest identity with PtALF5 at amino acid level but exhibited low similarity with most of other crustacean ALFs. Furthermore, different from the previously identified three SpALF homologs and most of other ALFs, SpALF4 had a low isoelectric point (pI) for the mature peptide and the LPS-binding domain with the values of 6.93 and 6.74, respectively. These results indicate that SpALF4 may be a unique ALF homolog with special biological function in the mud crab. Similar to the spatial structure of ALFPm3, SpALF4 contains three α-helices packed against a four-strand β-sheet, and an amphipathic loop formed by a disulphide bond between two conserved cysteine residues in LPS-binding domain. SpALF4, mainly distributed in hemocytes, could be upregulated by Vibrio harveyi, Staphylococcus aureus, or white spot syndrome virus. Recombinant SpALF4 could inhibit the growth of Gram-negative bacteria (V. harveyi, Vibrio anguillarum, Vibrio alginolyticus, Aeromonas hydrophila, Pseudomonas putida), Gram-positive bacteria (S. aureus and Bacillus megaterium), and a fungus Candida albicans to varying degrees. Further study showed that it could also bind to all the aforementioned microorganisms except S. aureus. These results demonstrate that SpALF4 is a unique ALF homolog with potent antimicrobial activity against bacteria and fungi. This characteristic suggests SpALF4 plays an essential function in immune defense against pathogen invasion in mud crab. •This study identified and characterized a novel ALF homolog (SpALF4) from mud crab.•Different from the most ALFs, SpALF4 had a low pI for the LPS-binding domain.•SpALF4 could inhibit the growth of all tested bacteria and fungi to varying degrees.•SpALF4 could also bind to all the tested microorganisms except Staphylococcus aureus.•SpALF4 is a unique ALF with potent antimicrobial activity against bacteria and fungi.