SpALF4: a newly identified anti-lipopolysaccharide factor from the mud crab Scylla paramamosain with broad spectrum antimicrobial activity.
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.
East China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Shanghai 200090, China; Key Laboratory of East China Sea and Oceanic Fishery Resources Exploitation, Ministry of Agriculture, Shanghai 200090, China.
Lei
L
Zhu
School of Life Sciences, Shandong University, Jinan, Shandong 250100, China.
Jiang-Feng
JF
Lan
East China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Shanghai 200090, China; Key Laboratory of East China Sea and Oceanic Fishery Resources Exploitation, Ministry of Agriculture, Shanghai 200090, China.
Yan-Qing
YQ
Huang
College of Fisheries, Henan Normal University, Xinxiang, Henan 453007, China.
Chao
C
Zhang
East China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Shanghai 200090, China; Key Laboratory of East China Sea and Oceanic Fishery Resources Exploitation, Ministry of Agriculture, Shanghai 200090, China.
Jun-Fang
JF
Zhou
East China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Shanghai 200090, China; Key Laboratory of East China Sea and Oceanic Fishery Resources Exploitation, Ministry of Agriculture, Shanghai 200090, China. Electronic address: fwenhong@163.com.
Wen-Hong
WH
Fang
East China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Shanghai 200090, China; Key Laboratory of East China Sea and Oceanic Fishery Resources Exploitation, Ministry of Agriculture, Shanghai 200090, China.
Xiao-Juan
XJ
Yao
East China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Shanghai 200090, China; Key Laboratory of East China Sea and Oceanic Fishery Resources Exploitation, Ministry of Agriculture, Shanghai 200090, China.
Hao
H
Wang
East China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Shanghai 200090, China; Key Laboratory of East China Sea and Oceanic Fishery Resources Exploitation, Ministry of Agriculture, Shanghai 200090, China. Electronic address: lixin8687@163.com.
Xin-Cang
XC
Li
D000595:Amino Acid Sequence; D000818:Animals; D023181:Antimicrobial Cationic Peptides / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D001483:Base Sequence; D003386:Brachyura / Q000235:genetics / Q000276:immunology*; D003001:Cloning, Molecular; D006090:Gram-Negative Bacteria / Q000276:immunology*; D006094:Gram-Positive Bacteria / Q000276:immunology*; D057927:Hydrophobic and Hydrophilic Interactions; D007526:Isoelectric Point; D008826:Microbial Sensitivity Tests; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010802:Phylogeny; D012313:RNA / Q000737:chemistry / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D013211:Staphylococcus aureus / Q000276:immunology*
D023181:Antimicrobial Cationic Peptides; D011994:Recombinant Proteins; D012313:RNA
Antimicrobial activity; Binding activity; Expression pattern; SpALF4