DNA binding activity of the baculovirus late expression factor PP31

Abstract

PP31 is a baculovirus protein that is essential for viral late gene expression. To study the role of PP31 in late transcription in vitro, it was purified from infected insect cells. A combination of heparin affinity, cation exchange chromatography, and gel filtration was used to purify native non-tagged protein. Nearly 5 mg of PP31 was obtained from 95 mg of nuclear extract confirming that PP31 is an abundant viral protein. DNA binding assays revealed that PP31 binds to single-stranded and double-stranded DNA with equal affinities. Addition of PP31 to in vitro transcription assays with purified baculovirus RNA polymerase resulted in a strong inhibition of transcription. This indicates that the viral RNA polymerase was not able to displace PP31, and suggests that other late expression factors may function to help RNA polymerase bind to PP31-coated templates.