Molecular cloning, expression and characterization of a functional GSTmu class from the cattle tick Boophilus annulatus.

摘要: Abstract A full-length cDNA of a glutathione S -transferase (GST) was cloned from library the local Egyptian cattle tick Boophilus annulatus . The 672 bp fragment sequenced and showed an open reading frame encoding protein 223 amino acids. Comparison deduced acid sequence with GSTs other species revealed that is closely related to mammalian mu-class GST. gene expressed in E. coli under T7 promotor pET-30b vector, purified native conditions. enzyme appeared as single band on 12% SDS-PAGE has molecular weight 30.8 kDa including histidine tag vector. assayed upon chromogenic substrate 1-chloro-2,4-dinitrobenzene (CDNB) recombinant high level activity even presence β-galactosidase region its 5′ end maximum at pH 7.5. K m values for CDNB GSH were 0.57 0.79 mM, respectively. over rBaGST toward (121 units/mg protein) less DCNB (29.3 units/mg protein). exhibited peroxidatic cumene hydroperoxide sharing this property belonging GST α class. I 50 cibacron blue bromosulfophthalein 0.22 8.45 μM, respectively, GSTmu Immunoblotting molecule B. extracts; whole tick, larvae, gut, salivary gland ovary. Homologues also detected Hyalomma dromedarii Rhipicephalus sp. while Ornithodoros moubata , homologue could not be detected.