Characterisation of a Plancitoxin-1-Like DNase II Gene in Trichinella spiralis

Background: deoxyribonuclease ii (dnase ii) is a well-known acidic endonuclease that catalyses the degradation of dna into oligonucleotides. only one or a few genes encoding dnase ii have been observed in the genomes of many species. 125 dnase ii-like protein family genes were predicted in the trichinella spiralis (t. spiralis) genome; however,none have been confirmed. dnase ii is a monomeric nuclease that contains two copies of a variant hkd motif in the n- and c-termini. of these 125 genes,only plancitoxin-1 (1095 bp,genbank accession no. xm_003370715.1) contains the hkd motif in its c-terminus domain. methodology/principal findings: in this study,we cloned and characterised the plancitoxin-1 gene. however,the sequences of plancitoxin-1 cloned from t. spiralis were shorter than the predicted sequences in genbank. intriguingly,there were two hkd motifs in the n- and c-termini in the cloned sequences. therefore,the gene with shorter sequences was named after plancitoxin-1-like (ts-pt,885 bp) and has been deposited in genbank under accession number kf984291. the recombinant protein (rts-pt) was expressed in a prokaryotic expression system and purified by nickel affinity chromatography. western blot analysis showed that rts-pt was recognised by serum from t. spiralis-infected mice; the anti-rts-pt serum recognised crude antigens but not es antigens. the ts-pt gene was examined at all t. spiralis developmental stages by real-time quantitative pcr. immunolocalisation analysis showed that ts-pt was distributed throughout newborn larvae (nbl),the tegument of adults (ad) and muscle larvae (ml). as demonstrated by dnase zymography,the expressed proteins displayed cation-independent dnase activity. rts-pt had a narrow optimum ph range in slightly acidic conditions (ph 4 and ph 5),and its optimum temperature was 25°c,30°c,and 37°c. conclusions: this study indicated that ts-pt was classified as a somatic protein in different t. spiralis developmental stages,and demonstrated for the first time that an expressed dnase ii protein from t. spiralis had nuclease activity. © 2014 liao et al.