Purification and characterization of a lectin from the white shrimp Litopenaeus setiferus (Crustacea decapoda) hemolymph
Purification and characterization of a lectin from the white shrimp Litopenaeus setiferus (Crustacea decapoda) hemolymph
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DOI：
10.1016/j.bbagen.2005.04.014
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发表时间：
2005-06-20
影响因子：
3
通讯作者：
Zenteno, E
中科院分区：
文献类型：
--
作者：
Alpuche, J;Pereyra, A;Zenteno, E
用戊二醛固定的兔红细胞基质亲和层析法从对虾血淋巴中纯化出291 kDa的凝集素(LSL)。LSL是一个由两个80 kDa和两个52 kDa亚基组成的杂四聚体，没有共价式碳水化合物，主要由天冬氨酸和谷氨酸、甘氨酸和丙氨酸组成，蛋氨酸和半胱氨酸含量相对较低。Edman降解表明，80 kDa亚基的NH2末端由DASNAQKQHDVNFLL组成，而52 kDa亚基的NH2末端被阻断。用MALDI-TOF从各亚基中预测LSL的多肽质量指纹图谱，结果表明各亚基与凡纳滨对虾血蓝蛋白前体的同源性分别为23%和22%。圆二色谱分析表明，β折叠和α螺旋的含量分别为52.7%和6.1%。LSL凝集效价较高的豚鼠、小鼠和兔红细胞，其活性依赖于二价阳离子。N-乙酰化糖，如GlcNAc、GaINAc和NeuAc是最有效的LSL血凝活性抑制剂。唾液酸化的O-糖基化蛋白，如牛颌下腺粘蛋白、人IgA和胎球蛋白，比唾液酸化的N-糖基化蛋白，如人乳粘蛋白、免疫球蛋白、转铁蛋白和乳铁蛋白具有更强的抑制活性。红细胞或抑制性糖蛋白的缺失消除了它们与LSL的结合能力，证实了唾液酸与LSL-配体相互作用的相关性。(C)2005 Elsevier B.V.保留所有权利。
A 291-kDa lectin (LsL) was purified from the hemolymph of the white shrimp Litopenaeus setiferus by affinity chromatography on glutaraldehyde-fixed stroma from rabbit erythrocytes. LsL is a heterotetramer of two 80-kDa and two 52-kDa subunits, with no covalently-liked carbohydrate, and mainly composed by aspartic and glutamic acids, glycine and alanine, with relatively lower methionine and cysteine contents. Edman degradation indicated that the NH2-terminal of the 80-kDa subunit is composed DASNAQKQHDVNFLL, whereas the NH2-terminal of the 52-kDa subunit is blocked. The peptide mass fingerprint of LsL was predicted from tryptic peptides from each subunit by MALDI-TOF, and revealed that each subunit showed 23 and 22%, respectively, homology with the hemocyanin precursor from Litopenaeus vannamei. Circular dichroism analysis revealed beta sheet and alpha helix contents of 52.7 and 6.1%, respectively. LsL agglutinate at higher titers guinea pig, murine, and rabbit erythrocytes its activity is divalent cation-dependent. N-acetylated sugars, such as GlcNAc, GaINAc, and NeuAc, were the most effective inhibitors of the LsL hemagglutinating activity. Sialylated O-glycosylated proteins, such as bovine submaxillary gland mucin, human IgA, and fetuin, showed stronger inhibitory activity than sialylated N-glycosylated proteins, such as human orosomucoid, IgG, transferrin, and lactoferrin. Desialylation of erythrocytes or inhibitory glycoproteins abolished their capacity to bind LsL, confirming the relevance of sialic acid in LsL-ligand interactions. (c) 2005 Elsevier B.V. All rights reserved.