title: Characterization and functional analysis of Trichinella spiralis Nudix hydrolase.
abstract: Trichinella spiralis Nudix hydrolase (TsNd) was identified by screening a T7 phage display cDNA library from T. spiralis intestinal infective larvae (IIL), and vaccination of mice with recombinant TsNd protein (rTsNd) or TsNd DNA vaccine produced a partial protective immunity. The aim of this study was to identify the characteristics and biological functions of TsNd in the process of invasion and development of T. spiralis larvae. Transcription and expression of TsNd gene at all developmental stages of T. spiralis were observed by qPCR and immunofluorescent test (IFT). The rTsNd had the Nd enzymatic activity to dGTP, NAD, NADP and CoA. Its kinetic properties on the preferred substrate dGTP were calculated, and the Vmax, Km, and kcat/Km values at pH 8.0 were 3.19 μM min(-1) μg(-1), 370 μM, and 144 s(-1) M(-1), respectively, in reaction matrix containing 5 mM Zn(2+) and 2 mM DTT. The rTsNd was active from 25 °C to 50 °C, with optimal activity at 37 °C. rTsNd was able to bind specifically to mouse intestinal epithelial cells (IECs) and promoted the larval invasion of IECs, whereas anti-rTsNd antibodies inhibited the larval invasion of IECs in a dose-dependent manner. Anti-rTsNd antibodies could kill T. spiralis infective larvae by an ADCC-mediated mechanism. Our results showed that the rTsNd protein was able to interact with host IECs, had the Nudix hydrolasing activity and the enzymatic activity appeared to be essential indispensable for the T. spiralis larval invasion, development and survival in host.
authors.affiliation: Department of Parasitology, Medical College of Zhengzhou University, Zhengzhou 450052, PR China.
authors.forename: Shao Rong
authors.initials: SR
authors.lastname: Long
authors.affiliation: Department of Parasitology, Medical College of Zhengzhou University, Zhengzhou 450052, PR China. Electronic address: wangzq@zzu.edu.cn.
authors.forename: Zhong Quan
authors.initials: ZQ
authors.lastname: Wang
authors.affiliation: Department of Parasitology, Medical College of Zhengzhou University, Zhengzhou 450052, PR China.
authors.forename: Peng
authors.initials: P
authors.lastname: Jiang
authors.affiliation: Department of Parasitology, Medical College of Zhengzhou University, Zhengzhou 450052, PR China.
authors.forename: Ruo Dan
authors.initials: RD
authors.lastname: Liu
authors.affiliation: Department of Parasitology, Medical College of Zhengzhou University, Zhengzhou 450052, PR China.
authors.forename: Xin
authors.initials: X
authors.lastname: Qi
authors.affiliation: Department of Parasitology, Medical College of Zhengzhou University, Zhengzhou 450052, PR China.
authors.forename: Pei
authors.initials: P
authors.lastname: Liu
authors.affiliation: Department of Parasitology, Medical College of Zhengzhou University, Zhengzhou 450052, PR China.
authors.forename: Hui Jun
authors.initials: HJ
authors.lastname: Ren
authors.affiliation: Department of Immunology, Massachusetts General Hospital and Harvard Medical School, Charlestown, MA 02129, USA.
authors.forename: Hai Ning
authors.initials: HN
authors.lastname: Shi
authors.affiliation: Department of Parasitology, Medical College of Zhengzhou University, Zhengzhou 450052, PR China. Electronic address: cuij@zzu.edu.cn.
authors.forename: Jing
authors.initials: J
authors.lastname: Cui
mesh_terms: D000818:Animals; D000909:Antibodies, Helminth / Q000276:immunology; D000920:Antibody-Dependent Cell Cytotoxicity; D015153:Blotting, Western; D004306:Dose-Response Relationship, Immunologic; D005260:Female; D005455:Fluorescent Antibody Technique; D015971:Gene Expression Regulation, Enzymologic; D006160:Guanosine Triphosphate / Q000378:metabolism; D006867:Hydrolases / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D006868:Hydrolysis; D007413:Intestinal Mucosa / Q000166:cytology / Q000469:parasitology; D008018:Life Cycle Stages / Q000235:genetics; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D018613:Microscopy, Confocal; D034421:Sus scrofa; D013552:Swine; D014158:Transcription, Genetic; D017160:Trichinella spiralis / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development / Q000276:immunology; D014235:Trichinellosis / Q000469:parasitology
chemical_list: D000909:Antibodies, Helminth; D006160:Guanosine Triphosphate; D006867:Hydrolases
keywords: Enzyme activity; Invasion; Molecular function; Nudix hydrolases; Trichinella spiralis