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A scallop C-type lectin from Argopecten irradians (AiCTL5) with activities of lipopolysaccharide binding and Gram-negative bacteria agglutination

Author:
Mu, Changkao, Song, Xiaoyan, Zhao, Jianmin, Wang, Lingling, Qiu, Limei, Zhang, Huan, Zhou, Zhi, Wang, Mengqiang, Song, Linsheng, Wang, Chunlin
Source:
Fish & shellfish immunology 2012 v.32 no.5 pp. 716-723
ISSN:
1050-4648
Subject:
Argopecten irradians, Bacillus thuringiensis, EDTA (chelating agent), Escherichia coli, Gram-negative bacteria, Gram-positive bacteria, Micrococcus luteus, Vibrio anguillarum, agglutination, amino acids, bacterial infections, carbohydrate binding, complementary DNA, erythrocytes, glycosylation, lectins, lipopolysaccharides, open reading frames, polypeptides, rabbits, scallops, signal peptide
Abstract:
C-type lectins are a family of calcium-dependent carbohydrate-binding proteins. In the present study, a C-type lectin (designated as AiCTL5) was identified and characterized from Argopecten irradians. The full-length cDNA of AiCTL5 was of 673 bp, containing a 5′ untranslated region (UTR) of 24 bp, a 3′ UTR of 130 bp with a poly (A) tail, and an open reading frame (ORF) of 519 bp encoding a polypeptide of 172 amino acids with a putative signal peptide of 17 amino acids. A C-type lectin-like domain (CRD) containing 6 conserved cysteines and a putative glycosylation sites were identified in the deduced amino acid sequence of AiCTL5. AiCTL5 shared 11%–27.5% identity with the previous reported C-type lectin from A. irradians. The cDNA fragment encoding the mature peptide of AiCTL5 was recombined into pET-21a (+) with a C-terminal hexa-histidine tag fused in-frame, and expressed in Escherichia coli Origami (DE3). The recombinant AiCTL5 (rAiCTL5) agglutinated Gram-negative E. coli TOP10F′ and Listonella anguillarum, but did not agglutinate Gram-positive bacteria Bacillus thuringiensis and Micrococcus luteus, and the agglutination could be inhibited by EDTA, indicating that AiCTL5 was a Ca²⁺-dependent lectin. rAiCTL5 exhibited a significantly strong activity to bind LPS from E. coli, which conformed to the agglutinating activity toward Gram-negative bacteria. Moreover, rAiCTL5 also agglutinated rabbit erythrocytes. These results indicated that AiCTL5 could function as a pattern recognition receptor to protect bay scallop from Gram-negative bacterial infection, and also provide evidence to understand the structural and functional diverse of lectin.
Agid:
829000