FUNCTIONAL EXPRESSION OF THE BOVINE HERPESVIRUS-1 ALKALINE DEOXYRIBONUCLEASE (UL12) IN ESCHERICHIA-COLI

Sequence analysis within the unique long segment of the bovine herpesv irus 1 (BHV-1; infectious bovine rhinotracheitis virus) genome identif ied an open reading frame whose deduced protein product of 487 amino a cids exhibited homology to alkaline deoxyribonucleases (DNases) of oth er herpesviruses. To determine this BHV-1 gene product has nuclease ac tivity, the gene designated UL12 was inserted into the vector pET-28a( +) and expressed in Escherichia coli as an oligohistidine-tagged prote in. Upon induction with isopropyl beta-D-thiogalactopyranoside E. coli BL21 (DE3) [pLysS] cells carrying this recombinant plasmid produced a 57-kDa protein, the molecular mass of which was in accordance with th e prediction from the DNA sequence. The recombinant UL12 protein purif ied by nickel-chelating affinity chromatography exhibited both exonucl ease and endonuclease activity, each with an alkaline pH optimum.