title: Molecular cloning and functional characterization of theta class glutathione S-transferase from Apostichopus japonicus.
abstract: Glutathione S-transferases (GSTs) are the superfamily of multifunctional detoxification isoenzymes and play crucial roles in innate immunity. In the present study, a theta class GST homology was identified from A. japonicus (designated as AjGST-θ) by RACE approaches. The full-length cDNA of AjGST-θ was of 1013 bp encoded a cytosolic protein of 231 amino acids residues. Structural analysis revealed that AjGST-θ processed the characteristic N-terminal GSH-binding site (G-site) and the C-terminal hydrophobic substrate binding site (H-site). Multiple sequence alignment and phylogenetic analysis together supported that AjGST-θ belonged to a new member of theta class GST protein subfamily. Spatial expression analysis revealed that AjGST-θ was ubiquitously expressed in all examined tissues with the larger magnitude in intestine. The Vibrio splendidus challenge in vivo and LPS stimulation in vitro could both significantly up-regulate the mRNA expression of AjGST-θ when compared with control group. The recombinant protein was expressed in Escherichia coli and the purified AjGST-θ showed high activity with GST substrate. Meantime, disc diffusion assay showed that recombinant AjGST-θ protein could markedly improve bacterial growth under Cumene hydroperoxide exposure. More importantly, the recombinant AjGST-θ could effectively prevent primary coelomocytes apoptosis after LPS exposure. Our present findings suggested that AjGST-θ might play significantly roles in the modulation of immune response and protect cells from pathogens infection in A. japonicus.
authors.affiliation: School of Marine Sciences, Ningbo University, Ningbo, 315211, PR China.
authors.forename: Yina
authors.initials: Y
authors.lastname: Shao
authors.affiliation: School of Marine Sciences, Ningbo University, Ningbo, 315211, PR China.
authors.forename: Zhimeng
authors.initials: Z
authors.lastname: Lv
authors.affiliation: School of Marine Sciences, Ningbo University, Ningbo, 315211, PR China. Electronic address: lichenghua@nbu.edu.cn.
authors.forename: Chenghua
authors.initials: C
authors.lastname: Li
authors.affiliation: School of Marine Sciences, Ningbo University, Ningbo, 315211, PR China.
authors.forename: Weiwei
authors.initials: W
authors.lastname: Zhang
authors.affiliation: School of Marine Sciences, Ningbo University, Ningbo, 315211, PR China.
authors.forename: Xuemei
authors.initials: X
authors.lastname: Duan
authors.affiliation: School of Marine Sciences, Ningbo University, Ningbo, 315211, PR China.
authors.forename: Qiongfen
authors.initials: Q
authors.lastname: Qiu
authors.affiliation: School of Marine Sciences, Ningbo University, Ningbo, 315211, PR China.
authors.forename: Chunhua
authors.initials: C
authors.lastname: Jin
authors.affiliation: School of Marine Sciences, Ningbo University, Ningbo, 315211, PR China.
authors.forename: Xuelin
authors.initials: X
authors.lastname: Zhao
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis; D001483:Base Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D005982:Glutathione Transferase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D007113:Immunity, Innate; D008070:Lipopolysaccharides / Q000494:pharmacology; D010802:Phylogeny; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D047528:Stichopus / Q000145:classification / Q000201:enzymology* / Q000235:genetics* / Q000382:microbiology; D014733:Vibrio / Q000502:physiology
chemical_list: D018076:DNA, Complementary; D008070:Lipopolysaccharides; D012333:RNA, Messenger; D011994:Recombinant Proteins; D005982:Glutathione Transferase
keywords: Apostichopus japonicus; Gene expression; Glutathione S-transferase; Immune response