Nucleotides within both proximal and distal parts of the consensus sequence are important for specific DNA recognition by the herpes simplex virus regulatory protein ICP4.
The herpes simplex virus type 1 regulatory protein ICP4 is a sequence specific DNA binding protein which associates with a number of different sites, some of which include the consensus ATCGTCnnnnYCGRC. In order to investigate the involvement in DNA binding of conserved bases within the consensus, we have synthesised a family of mutant oligonucleotides and tested their ability to form a complex with ICP4. We have also compared the binding specificities of bacterially expressed fragments of ICP4 which include the DNA binding domain. Mutation of most (but not all) bases in the proximal part of the consensus greatly reduced binding by ICP4, as did a mutation affecting the distal part. Most (but not all) G residues identified in methylation interference assays were required for efficient binding. While a bacterially expressed ICP4 peptide encompassing amino acid residues 252-523 bound to DNA with a specificity similar to that of the whole protein, a shorter protein (residues 275-523) had a slightly relaxed DNA binding specificity.
Pizer
L I
LI
Department of Microbiology/Immunology, University of Colorado Health Sciences Center, Denver 80262.
Everett
R D
RD
Tedder
D G
DG
Elliott
M
M
Litman
B
B
D001483:Base Sequence; D001665:Binding Sites; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000502:physiology*; D006367:HeLa Cells; D017874:Immediate-Early Proteins; D008745:Methylation; D008969:Molecular Sequence Data; D009841:Oligonucleotides / Q000737:chemistry; D011994:Recombinant Proteins / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid; D018139:Simplexvirus / Q000235:genetics*; D013329:Structure-Activity Relationship; D014759:Viral Envelope Proteins / Q000235:genetics; D054334:Viral Regulatory and Accessory Proteins / Q000502:physiology*
D004268:DNA-Binding Proteins; D017874:Immediate-Early Proteins; D009841:Oligonucleotides; D011994:Recombinant Proteins; D014759:Viral Envelope Proteins; D054334:Viral Regulatory and Accessory Proteins; C039447:glycoprotein D, Human herpesvirus 1; C048925:herpes simplex virus, type 1 protein ICP4