Identification of a DNA binding domain in simian virus 40 capsid proteins Vp2 and Vp3.
We have identified both biochemically and genetically a protein domain within the simian virus 40 virion protein Vp3, and within Vp2 since its carboxyl two-thirds are identical to the full-length Vp3, that binds DNA in a sequence nonspecific manner. Both the Vp2 and Vp3 (Vp2/3) components of SV40 and mutant SV40(202T) bound either SV40 or pBR322 DNA equally well. Wild type and mutant Vp2/3 proteins, expressed as fusion proteins with glutathione S-transferase (GST), were tested for their ability to bind DNA. GST-Vp3 bound DNA at physiological salt concentrations with an apparent Kd of 2.5 x 10(-8) M and also bound RNA with 4-fold higher affinity. Over 90% of the nucleic acid binding, and all of the activity, was lost upon removal of the carboxyl-terminal 13 and 35 residues, respectively. The DNA binding domain was shown to be distinct and separable from the Vp2/3 nuclear transport signal since mutations within the nuclear transport signal that reduce or abolish nuclear localization of Vp2/3 had no effect on the DNA binding activity of mutant Vp2/3 fusion proteins. The carboxyl-terminal 40 residues of Vp2/3 in the form of a beta-galactosidase fusion protein, F6, are sufficient for DNA binding and may cause compaction of the DNA. The significance of this DNA binding and possible compaction are discussed in relation to the assembly of virion particles.
Clever
J
J
Department of Biology, University of California, Los Angeles 90024.
Dean
D A
DA
Kasamatsu
H
H
D000818:Animals; D001665:Binding Sites; D015139:Blotting, Southern; D015153:Blotting, Western; D002213:Capsid / Q000378:metabolism*; D036022:Capsid Proteins; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D004274:DNA, Recombinant / Q000378:metabolism; D004279:DNA, Viral / Q000378:metabolism*; D005982:Glutathione Transferase / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013539:Simian virus 40 / Q000235:genetics* / Q000378:metabolism; D012996:Solutions; D014771:Virion / Q000378:metabolism; D001616:beta-Galactosidase / Q000378:metabolism
D036022:Capsid Proteins; D004274:DNA, Recombinant; D004279:DNA, Viral; D011993:Recombinant Fusion Proteins; D012996:Solutions; C442543:VP2 protein, Polyomavirus; D005982:Glutathione Transferase; D001616:beta-Galactosidase