Evidence that vaccinia virulence factor E3L binds to Z-DNA in vivo: Implications for development of a therapy for poxvirus infection.
The E3L gene product found in all poxviruses is required for the lethality of mice in vaccinia virus infection. Both the C-terminal region, consisting of a double-stranded RNA-binding motif, and the N-terminal region (vZ(E3L)), which is similar to the Zalpha family of Z-DNA-binding proteins, are required for infection. It has recently been demonstrated that the function of the N-terminal domain depends on its ability to bind Z-DNA; Z-DNA-binding domains from unrelated mammalian proteins fully complement an N-terminal deletion of E3L. Mutations that decrease affinity for Z-DNA have similar effects in decreasing pathogenicity. Compounds that block the Z-DNA-binding activity of E3L may also limit infection by the poxvirus. Here we show both an in vitro and an in vivo assay with the potential to be used in screening for such compounds. Using a conformation-specific yeast one-hybrid assay, we compared the results for Z-DNA binding of vZ(E3L) with those for human Zbeta(ADAR1), a peptide that has similarity to the Zalpha motif but does not bind Z-DNA, and with a mutant of hZbeta(ADAR1), which binds Z-DNA. The results suggest that this system can be used for high-throughput screening.
Kim
Yang-Gyun
YG
Department of Biology, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Cambridge, MA 02139, USA.
Lowenhaupt
Ky
K
Oh
Doo-Byoung
DB
Kim
Kyeong Kyu
KK
Rich
Alexander
A
D000595:Amino Acid Sequence; D000818:Animals; D002942:Circular Dichroism; D043542:DNA, Z-Form; D051379:Mice; D008969:Molecular Sequence Data; D011213:Poxviridae Infections / Q000628:therapy*; D011485:Protein Binding; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D020798:Two-Hybrid System Techniques; D014764:Viral Proteins / Q000737:chemistry / Q000378:metabolism*
D043542:DNA, Z-Form; C081374:E3L protein, Vaccinia virus; D016601:RNA-Binding Proteins; D014764:Viral Proteins