Expression and functional validation of Bombyx mori nucleopolyhedrovirus ORF29, a conserved Nudix motif protein.

Our previous study showed that Bombyx mori nucleopolyhedrovirus (BmNPV) orf29 encodes a 26 kDa protein expressed in the early stage of infection cycle. BmNPV ORF29, contains a conserved motif of Nudix (nucleotide diphosphate X) superfamily. It has the highest homology with ADP-ribose pyrophosphatase (ADPRase), a subfamily of Nudix pyrophosphatase. In this work, we purified the recombinant BmNPV ORF29 in Escherichia coli by metal chelating affinity chromatography. The amino acid sequence of recombinant protein was confirmed by mass spectroscopic analysis and found that the purified protein could be able to catalyze the breakdown of ADP-ribose to AMP and ribose 5-phosphate, with Km and Kcat values of 182 μmol/l and 5.3 s-1 respectively. The optimal activity was at alkaline pH (8.5) with Mg2+ (0.5-mmol/l) ions as the cofactor.