Molecular expression and characterization of GCP7 gene of Haemonchus contortus

Abstract

Haemonchus contortus is a highly pathogenic and most economically important parasite of sheep and goats worldwide. The cysteine proteases from H. contortus are prime targets for vaccine development. In the present communication we report the molecular expression and characterization of cathepsin B-like cysteine protease, GCP7 gene of H. contortus to study its efficiency as target protein for immunoprophylaxis against haemonchosis in sheep and goats. The complete ORF of GCP7 gene, devoid of the signal sequence, was amplified by RT-PCR from mRNA isolated from H. contortus and was cloned initially into pTZ57R/T cloning vector and then sub-cloned in the pET32a(+) expression vector to produce GCP7 antigen. The nucleotide and deduced amino acid sequence of the GCP7 was aligned against the related sequences of H. contortus available in public domain for in silico analysis by DNA STAR and MEGA version 4.0 softwares. The nucleotide sequence revealed that the GCP7 gene of H.contortus (Indian isolate) encodes 324 amino acids (devoid of signal sequence) and its nucleotide sequence had 95.9% to 99.4% sequence homology with that of U.S.A. and previously published Indian isolates. A high level expression of recombinant (r) GCP7 protein was observed in the molecular range (Mr) of 55 kDa. The rGCP7 protein was confirmed by its specific immunoreactivity against known reference positive sheep sera.