title: Lipopolysaccharide and β-1,3-glucan-binding protein (LGBP) bind to seaweed polysaccharides and activate the prophenoloxidase system in white shrimp Litopenaeus vannamei.
abstract: Lipopolysaccharide and β-1,3-glucan-binding protein (LGBP), important pattern recognition proteins (PRPs), recognize lipopolysaccharide (LPS) and β-1,3-glucan (βG), known as pathogen-associated molecular patterns (PAMPs), and subsequently trigger innate immunity. Several seaweed polysaccharides and seaweed extracts increase immune parameters and resistance to pathogens. Here, we constructed the expression vector pET28b-LvLGBP and transferred it into Escherichia coli BL21 (DE3) for protein expression and to produce the recombinant protein LGBP (rLvLGBP) in white shrimp Litopenaeus vannamei. We examined the binding of rLvLGBP with seaweed-derived polysaccharides including alginate, carrageenan, fucoidan, laminarin, Gracilaria tenuistipitata extract (GTE), and Sargassum duplicatum extract (SDE), and examined the phenoloxidase activity of shrimp haemocytes incubated with a mixture of rLvLGBP and each polysaccharide. We also examined the binding of rLvLGBP with LPS and βG, and the phenoloxidase activity of shrimp haemocytes incubated with a mixture of rLvLGBP and LPS (rLvLGBP-LPS) or a mixture of rLvLGBP and βG (rLvLGBP-βG). An ELISA binding assay indicated that rLvLGBP binds to LPS, βG, alginate, carrageenan, fucoidan, laminarin, GTE, and SDE with dissociation constants of 0.1138-0.1770 μM. Furthermore, our results also indicated that the phenoloxidase activity of shrimp haemocytes incubated with a mixture of rLvLGBP and LPS, βG, alginate, carrageenan, fucoidan, laminarin, GTE, and SDE significantly increased by 328%, 172%, 200%, 213%, 197%, 194%, 191%, and 197%, respectively compared to controls (cacodylate buffer). We conclude that LvLGBP functions as a PRP, recognizes and binds to LPS, βG, alginate, carrageenan, fucoidan, laminarin, GTE, and SDE, and subsequently leads to activating innate immunity in shrimp.
authors.affiliation: Department of Aquaculture, College of Life Sciences, Center of the Excellence for the Oceans, National Taiwan Ocean University, Keelung, 20224, Taiwan.
authors.forename: Yu-Yuan
authors.initials: YY
authors.lastname: Chen
authors.affiliation: Department of Aquaculture, College of Life Sciences, Center of the Excellence for the Oceans, National Taiwan Ocean University, Keelung, 20224, Taiwan. Electronic address: jcchen@mail.ntou.edu.tw.
authors.forename: Jiann-Chu
authors.initials: JC
authors.lastname: Chen
authors.affiliation: Department of Aquaculture, College of Life Sciences, Center of the Excellence for the Oceans, National Taiwan Ocean University, Keelung, 20224, Taiwan.
authors.forename: Yi-Hsuan
authors.initials: YH
authors.lastname: Kuo
authors.affiliation: Department of Aquaculture, College of Life Sciences, Center of the Excellence for the Oceans, National Taiwan Ocean University, Keelung, 20224, Taiwan.
authors.forename: Yong-Chin
authors.initials: YC
authors.lastname: Lin
authors.affiliation: Department of Aquaculture, College of Life Sciences, Center of the Excellence for the Oceans, National Taiwan Ocean University, Keelung, 20224, Taiwan.
authors.forename: Yu-Hsuan
authors.initials: YH
authors.lastname: Chang
authors.affiliation: Department of Aquaculture, College of Life Sciences, Center of the Excellence for the Oceans, National Taiwan Ocean University, Keelung, 20224, Taiwan.
authors.forename: Hong-Yi
authors.initials: HY
authors.lastname: Gong
authors.affiliation: Department of Aquaculture, College of Life Sciences, Center of the Excellence for the Oceans, National Taiwan Ocean University, Keelung, 20224, Taiwan.
authors.forename: Chien-Lun
authors.initials: CL
authors.lastname: Huang
mesh_terms: D000818:Animals; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D004156:Catechol Oxidase / Q000378:metabolism*; D004792:Enzyme Precursors / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D044724:Gracilaria / Q000276:immunology; D006434:Hemocytes / Q000502:physiology*; D007113:Immunity, Innate; D037102:Lectins / Q000235:genetics / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000276:immunology; D033561:Penaeidae / Q000276:immunology*; D010936:Plant Extracts / Q000276:immunology; D051192:Receptors, Pattern Recognition / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics; D044647:Sargassum / Q000276:immunology; D047071:beta-Glucans / Q000378:metabolism
chemical_list: D002352:Carrier Proteins; D004792:Enzyme Precursors; D037102:Lectins; D008070:Lipopolysaccharides; D010936:Plant Extracts; D051192:Receptors, Pattern Recognition; D011994:Recombinant Proteins; D047071:beta-Glucans; C037937:glucan-binding proteins; C033363:beta-1,3-glucan; C038451:pro-phenoloxidase; D004156:Catechol Oxidase
keywords: Lipopolysaccharide and β-1,3-glucan-binding protein (LGBP); Litopenaeus vannamei; Phenoloxidase; Recognition; Seaweed polysaccharide