Hepatitis B virus protein pX enhances the monomer assembly pathway of bZIP.DNA complexes.
The rapid and correct assembly of dimeric transcription factors on target DNA is essential for accurate transcriptional regulation. Here we ask how a viral accessory factor, hepatitis B virus X protein (pX), influences the rate and identity of the assembly pathway followed by members of the basic region leucine zipper (bZIP) transcription factor family. A combination of fluorescence polarization and fluorescence resonance energy transfer (FRET) experiments demonstrates unequivocally that pX does not increase the concentration of properly folded bZIP dimers in solution. Rather, fluorescence polarization and gel mobility shift experiments reveal that pX interacts directly with the basic-spacer segment of the bZIP peptide and stabilizes the complex composed of this monomer and target DNA. By stabilizing the intermediate formed along the monomer assembly pathway but not the one formed along the dimer pathway, pX enhances the equilibrium stability of a bZIP.DNA complex without changing the molecular mechanism used for complexation. Additional experiments reveal that pX decreases the kinetic specificity of certain bZIP proteins. To the extent that it is reflected at the transcriptional level, this loss in specificity could have far-reaching consequences for the host cell.
Department of Chemistry, Yale University, New Haven, Connecticut 06511, USA.
T L
TL
Schneider
A
A
Schepartz
D000595:Amino Acid Sequence; D050976:Basic-Leucine Zipper Transcription Factors; D004268:DNA-Binding Proteins / Q000378:metabolism*; D019281:Dimerization; D004591:Electrophoresis, Polyacrylamide Gel; D004735:Energy Transfer; D005452:Fluoresceins / Q000378:metabolism; D005454:Fluorescence Polarization; D005456:Fluorescent Dyes / Q000378:metabolism; D051790:G-Box Binding Factors; D006515:Hepatitis B virus / Q000502:physiology*; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D009841:Oligonucleotides / Q000378:metabolism; D011485:Protein Binding; D012235:Rhodamines / Q000378:metabolism; D017384:Sequence Deletion; D013050:Spectrometry, Fluorescence; D015534:Trans-Activators / Q000138:chemical synthesis / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D014157:Transcription Factors / Q000378:metabolism*; D054334:Viral Regulatory and Accessory Proteins
D050976:Basic-Leucine Zipper Transcription Factors; D004268:DNA-Binding Proteins; D005452:Fluoresceins; D005456:Fluorescent Dyes; D051790:G-Box Binding Factors; D009841:Oligonucleotides; D012235:Rhodamines; D015534:Trans-Activators; D014157:Transcription Factors; D054334:Viral Regulatory and Accessory Proteins; C057628:hepatitis B virus X protein; C015296:5-iodoacetamidofluorescein; C034996:tetramethylrhodamine iodoacetamide