Localization of a 34-amino-acid segment implicated in dimerization of the herpes simplex virus type 1 ICP4 polypeptide by a dimerization trap.
The herpes simplex virus type 1 immediate-early protein ICP4 plays an essential role in the regulation of the expression of all viral genes. It is the major trans activator of early and late genes and also has a negative regulatory effect on immediate-early gene transcription. ICP4 is a sequence-specific DNA-binding protein and has always been purified in a dimeric form. The part of the protein that consists of the entire highly conserved region 2 and of the distal portion of region 1 retains the ability to specifically associate with DNA and to form homodimers in solution. In an attempt to map the dimerization domain of ICP4, we used a dimerization trap assay, in which we screened deletion fragments of this 217-amino-acid stretch for sequences that could confer dimerization properties on a heterologous cellular transcription factor (LFB1), which binds to its cognate DNA sequence only as a dimer. The analysis of these chimeric proteins expressed in vitro ultimately identified a stretch of 34 amino acids (343 to 376) that could still confer DNA-binding activity on the LFB1 reporter protein and thus apparently contained the ICP4 dimerization motif. Consistent with this result, a truncated ICP4 protein containing amino acids 343 to 490, in spite of the complete loss of DNA-binding activity, appeared to retain the capacity to form a heterodimer with a longer ICP4 peptide after coexpression in an in vitro translation system.
Gallinari
P
P
Istituto di Richerche di Biologia Molecolare P. Angeletti, Pomezia, Italy.
Wiebauer
K
K
Nardi
M C
MC
Jiricny
J
J
D000595:Amino Acid Sequence; D001483:Base Sequence; D004279:DNA, Viral / Q000235:genetics; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D018259:Herpesvirus 1, Human / Q000737:chemistry / Q000235:genetics*; D017874:Immediate-Early Proteins / Q000737:chemistry / Q000235:genetics*; D008969:Molecular Sequence Data; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics; D017384:Sequence Deletion; D015534:Trans-Activators / Q000737:chemistry / Q000235:genetics
D004279:DNA, Viral; D004268:DNA-Binding Proteins; D017874:Immediate-Early Proteins; D011993:Recombinant Fusion Proteins; D015534:Trans-Activators; C048925:herpes simplex virus, type 1 protein ICP4