DNA binding activity of the baculovirus late expression factor PP31.
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.
Guarino
Linda A
LA
Department of Biochemistry and Biophysics, Texas A&M University, College Station, TX 77843-2128, USA. lguarino@tamu.edu
Mistretta
Toni-Ann
TA
Dong
Wen
W
D000818:Animals; D002478:Cells, Cultured; D004247:DNA / Q000378:metabolism*; D004277:DNA, Single-Stranded / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism*; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism; D015967:Gene Expression Regulation, Viral; D017924:Nucleopolyhedroviruses / Q000201:enzymology / Q000235:genetics / Q000378:metabolism; D010750:Phosphoproteins / Q000302:isolation & purification / Q000378:metabolism*; D018411:Spodoptera / Q000821:virology; D014158:Transcription, Genetic
D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D010750:Phosphoproteins; C078031:phosphoprotein pp31, Baculovirus; D004247:DNA; D012321:DNA-Directed RNA Polymerases