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

Abstract

Background Pyruvate 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 findings TsPKM 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. Conclusions TsPKM 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.

A reanalysis and integration of transcriptomics and proteomics datasets unveil novel drug targets for Mekong schistosomiasis

“…spiralis . Silencing TsPKM gene expression significantly impacted enzyme activity, larval molting, sugar metabolism, and overall parasite growth, suggesting TsPKM as a promising therapeutic target for trichinellosis treatment 36 . Due to its abundance in both genders, pyruvate kinase in S .…”

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

“…The soluble crude antigens of diverse T . spiralis stage worms (ML, IIL, AW and NBL) and their ES antigens were prepared as reported before [ 29 , 30 ].…”

A novel trypsin of Trichinella spiralis mediates larval invasion of gut epithelium via binding to PAR2 and activating ERK1/2 pathway

“…Finally, the bound proteins (protein A/G-rTsTryp-PAR2) were eluted from the beads, denatured by boiling for 5 min, and then separated by 12% SDS-PAGE and analyzed by western blotting, in which anti-rTsTryp antibody and anti-PAR2 antibody (1:1000; Abcam, UK) were used as the primary antibodies. The secondary antibody without light and heavy chains (1:5000, Abmart) was used to avoid IgG light and heavy chain pollution [50,51].…”