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Characterization of a Novel Glutamine Synthetase From Trichinella spiralis and Its Participation in Larval Acid Resistance, Molting, and Development.

ABSTRACT: Trichinella spiralis is a major foodborne parasite worldwide. After the encapsulated muscle larvae (ML) in meat are ingested, the ML are liberated in the stomach of the host and activated into intestinal infectious larvae (IIL), which develop into adult worm after molting four times. A novel glutamine synthetase (TsGS) was identified from T. spiralis IIL at 10 h post-infection, but its biological role in T. spiralis life cycle is not clear. The aim of this study was to investigate the biological characteristics of TsGS and its functions in larval acid resistance, molting, and development. TsGS has a glutamine synthetase (GS) catalytic domain. Complete TsGS sequence was cloned and expressed in Escherichia coli BL21. rTsGS has good immunogenicity. qPCR and Western blotting showed that TsGS was highly expressed at IIL stage, and immunofluorescence revealed that TsGS was principally localized at the cuticle and intrauterine embryos of this nematode. rTsGS has enzymatic activity of natural GS to hydrolyze the substrate (Glu, ATP, and NH₄ ⁺). Silencing of TsGS gene significantly reduced the IIL survival at pH 2.5, decreased the IIL burden, and impeded larval molting and development. The results demonstrated that TsGS participates in T. spiralis larval acid resistance, molting and development, and it might be a candidate vaccine target against Trichinella molting and development.

SUBMITTER: Zhuo TX

PROVIDER: S-EPMC8488193 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Characterization of a novel pyruvate kinase from Trichinella spiralis and its participation in sugar metabolism, larval molting and development.

Project description:BackgroundPyruvate kinase widely exists in many parasites and plays an important role in the energy production for the parasites. Pyruvate kinase might be a potential drug target for killing the parasites. The aim of the present study was to evaluate the biological characteristics and roles of T. spiralis pyruvate kinase M (TsPKM) in sugar metabolism, larval molting and development of T. spiralis.Methodology/principal findingsTsPKM has two functional domains of pyruvate kinase and the tertiary structure of TsPKM is tetramer which has the enzyme active site constituted by 8 amino-acid residues (Arg71, Asn73, Asp110, Phe241, Lys267, Glu269, Asp293 and Thr325). Recombinant TsPKM (rTsPKM) was expressed and purified. The rTsPKM had good immunogenicity. RT-PCR and Western blot showed that TsPKM was transcribed and expressed at various developmental stages in T. spiralis lifecycle. Immunofluorescence test showed that TsPKM was principally located in the cuticle, muscle, stichosome, intestine and the intrauterine embryos of female adults. rTsPKM catalyzed the reaction of phosphoenolpyruvate (PEP) and adenosine diphosphate (ADP) to produce pyruvic acid and adenosine triphosphate (ATP). TsPKM played an important role in the metabolism and energy production of T. spiralis. After silencing of TsPKM gene by specific dsRNA-TsPKM2, protein expression and enzyme activity of TsPKM decreased by 50.91 and 26.06%, respectively. After treatment with RNAi, natural TsPKM enzyme activity, larval molting, sugar metabolism, growth and development of T. spiralis were significantly reduced.ConclusionsTsPKM participates in the larval molting, sugar metabolism, growth and development of T. spiralis and it might be a candidate target of therapeutic drug of trichinellosis.

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Molecular characterization of Trichinella spiralis galectin and its participation in larval invasion of host's intestinal epithelial cells.

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Molecular characterization of a novel serine proteinase from Trichinella spiralis and its participation in larval invasion of gut epithelium.

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Characterization of a novel cysteine protease in Trichinella spiralis and its role in larval intrusion, development and fecundity.

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| S-EPMC8390047 | biostudies-literature

RNAi-mediated silencing of Trichinella spiralis glutaminase results in reduced muscle larval infectivity.

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| S-EPMC7992778 | biostudies-literature

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