Isolation, Characterization and Antibacterial Activity of Lectin ofCoelomic Fluid in Sipuncula Phasclosoma esculenta
CAI Binxin1, ZHOU Fengfang2, HUANG Weiqing1, WEI Qi1, DING Jianfa2
1. College of Marine Sciences, Ningde Normal University, Ningde 352100, China; 2. College of Biological Science and Engineering, Ningde Normal University, Ningde 352100, China
Abstract：Lectin wasisolated and purified in coelomic fluid of sipuncula Phascolosoma esculenta by precipitation and DEAE Sepharose Fast. Then characteristics and antibacterial activity were analyzed in the lectin under different temperature (30, 40, 50, 60 and 70 ℃), pH (3.0, 4.0, 5.0, 6.0, 7.0, 8.0, 9.0 and 10.0), metal ions (solution of divalent metal ions of BaCl2, MgSO4, FeCl2, ZnSO4, CaCl2, CuSO4 and MnCl2, repectively), and sugars (glucose, D-galactose, D-mannose, xylose, arabinose, rhamnose, lactose, sucrose, trehalose, N-acetylglucosamine, and N-acetyl D-galactosamine). The results showed that the activity of the lectin was high using 30%—40% ammonium sulfate-extraction, with four peaks (0.05×103, 3.43×103, 0.11×103 and 0.38×103 HU/mg, respectively) by DEAE Sepharose Fast chromatography and the stronger agglutination activity at the second peak, named as P. esculenta lectin. There was increase in specific activity to 18.08 fold, and only 0.82% of protein recovery rate. The P. esculenta lectin had relative molecula mass of about 35.5 ku, and the agglutination activity at pH from 7 to 8 and at below 40 ℃, and inhibited by D-galactose, lactose and N-acetyl-D-galactosamine. The agglutination activity was found to be promoted by Ca2+, increasing to 7.47×103HU/mg. The P. esculenta lectin showed high antimicrobial activity against Vibrio parahaemolyticus and Aeromonas hydrophila. The finding indicates that it shows high agglutinating activity, relatively stable properties, and inhibitory effects on V. parahaemolyticus and A. hydrophila.
[1] FEARON D T, LOCKSLEY R M. The instructive role of innate immunity in the acquired immune response[J].Science,1996,272(5258):50-54. [2] HOOPER C, DEBNATH P P, STENTIFORD G D, et al.Diseases of the giant river prawn Macrobrachium rosenbergii:a review for a growing industry[J].Reviews in Aquaculture,2023,15(2):738-758. [3] OLIVEIRA M V, OSTERNE V J S, LOSSIO C F, et al. Purification and characterization of a highly thermostable GlcNAc-binding lectin from Collaea speciosa seeds[J].International Journal of Biological Macromolecules,2021,193:1562-1571. [4] 李红岩,张祥敏,杨帅奇.海洋生物C型凝集素研究进展[J].中国海洋大学学报(自然科学版),2020,50(9):113-119. [5] ZENG B X, DONG T W, XIA Y T, et al.Characterization and comparison of two C-type lectins with novel key motifs from mud crab Scylla paramamosain[J].Journal of Oceanology and Limnology,2022,40(4):1628-1639. [6] ZHOU K M, QIN Y K, SONG Y, et al. A novel ig domain-containing C-type lectin triggers the intestine-hemocyte axis to regulate antibacterial immunity in crab[J].The Journal of Immunology,2022,208(10):2343-2362. [7] ZHANG Y, NI M Q, ZHANG P, et al. Identification and functional characterization of C-type lectins and crustins provide new insights into the immune response of Portunus trituberculatus[J].Fish & Shellfish Immunology,2022,129:170-181. [8] XIN Z Z, ZHANG X T, ZHOU M, et al. Differential molecular responses of hemolymph and hepatopancreas of swimming crab, Portunus trituberculatus, infected with Ameson portunus (Microsporidia)[J].Fish & Shellfish Immunology,2024,145:109324. [9] HUANG M M, WANG L L, ZHANG H, et al. The sequence variation and functional differentiation of CRDs in a scallop multiple CRDs containing lectin[J].Developmental & Comparative Immunology,2017,67:333-339. [10] YANG J L, HUANG M M, ZHANG H, et al. CfLec-3 from scallop:an entrance to non-self recognition mechanism of invertebrate C-type lectin[J].Scientific Reports,2015,5:10068. [11] MA T H, BENZIE J A H, HE J G, et al. PmLT, a C-type lectin specific to hepatopancreas is involved in the innate defense of the shrimp Penaeus monodon[J].Journal of Invertebrate Pathology,2008,99(3):332-341. [12] WANG H, XUE Z, LIU Z Q, et al. A novel C-type lectin from the sea cucumber Apostichopus japonicus (AjCTL-2) with preferential binding of d[J].Fish & Shellfish Immunology,2018,79:218-227. [13] 管华诗,王曙光.中华海洋本草-第3卷-海洋无脊椎动物药[M].上海:上海科学技术出版社,2009:78-90. [14] 戴聪杰,董乐,王芳.白玉蜗牛凝集素的分离纯化及凝集活性[J].泉州师范学院学报,2017,35(6):6-11. [15] 刘晓晨.鲟鱼体表黏液及其凝集素分离、鉴定的研究[D].大连:大连海洋大学,2022:22. [16] VARGIL A F, BAI S M M, MARY J V J, et al. Isolation, characterization and antimicrobial properties of hepatopancreas lectin of the freshwater crab Oziotelphusa naga[J].Protein Expression and Purification,2024,222:106536. [17] 蔡彬新,周逢芳,阮少江,等.养殖可口革囊星虫纤溶酶分离纯化及酶学特性研究[J].水产科学,2021,40(6):922-928. [18] 徐海圣,徐步进.中华绒螯蟹血清凝集素的分离纯化与性质研究[J].水产科学,2008,27(9):455-461. [19] HUANG Y, REN Q. Research progress in innate immunity of freshwater crustaceans[J].Developmental & Comparative Immunology,2020,104:103569. [20] IMAMICHI Y, YOKOYAMA Y. Purification and characterization of a lectin from the starfish Asterias amurensis[J].Fisheries Science,2013,79(6):1007-1013. [21] 梁青龙,苏鑫龙,戴聪杰.拟穴青蟹凝集素的活性及其部分性质[J].泉州师范学院学报,2012,30(2):72-75. [22] DILNA C, PRASANTH G K, GHUFRAN M S, et al. Purification and characterization of a hemocyanin with lectin-like activity isolated from the hemolymph of speckled shrimp, Metapenaeus monoceros[J].Biochimie,2023,206:36-48. [23] MIZGINA T O, CHIKALOVETS I V, BULANOVA T A, et al. New l-rhamnose-binding lectin from the bivalve Glycymeris yessoensis:purification, partial structural characterization and antibacterial activity[J].Marine Drugs,2024,22(1):27. [24] PALHARINI J G, RICHTER A C, SILVA M F, et al. Eutirucallin:a lectin with antitumor and antimicrobial properties[J].Frontiers in Cellular and Infection Microbiology,2017,7:136. [25] LEE C G, DA SILVA C A, DELA CRUZ C S, et al. Role of chitin and chitinase/chitinase-like proteins in inflammation, tissue remodeling, and injury[J].Annual Review of Physiology,2011,73:479-501. [26] POHLEVEN J, TRUKELJ B, KOS J. Afinity chromatography of lectins[M].Affinity Chromatography. InTech, 2012:49-74. [27] JIN X K, LI S, GUO X N, et al. Two antibacterial C-type lectins from crustacean, Eriocheir sinensis, stimulated cellular encapsulation in vitro[J].Developmental & Comparative Immunology,2013,41(4):544-552.
2026, Vol. 45 
