Antibody-promoted dimerization bypasses the regulation of DNA binding by the heme domain of the yeast transcriptional activator HAP1.
The yeast transcriptional activator HAP1 contains a DNA-binding domain homologous to the zinc finger of GAL4 and an adjacent regulatory domain that blocks DNA binding in the absence of the inducer heme. We show that short HAP1 fragments containing the zinc finger are unable to bind to DNA but can be rescued by antibody to the HAP1 zinc finger. These fragments are missing a coiled-coil sequence similar to that within the dimerization domain of GAL4 and dimerization domains of myosin heavy chain. We surmise that the antibody promotes DNA binding by bringing together two monomers. Interestingly, the antibody will also promote DNA binding of a larger HAP1 fragment containing the DNA-binding and the heme-regulatory domains. This suggests that the regulatory domain acts by preventing dimerization of HAP1 in the absence of heme. Consistent with this view is an in vivo assay that also reveals that heme promotes HAP1 dimerization in yeast cells.
Zhang
L
L
Department of Biology, Massachusetts Institute of Technology, Cambridge 02139.
Bermingham-McDonogh
O
O
Turcotte
B
B
Guarente
L
L
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies; D004268:DNA-Binding Proteins / Q000276:immunology / Q000378:metabolism*; D005656:Fungal Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D006418:Heme / Q000378:metabolism; D046911:Macromolecular Substances; D008961:Models, Structural; D008969:Molecular Sequence Data; D010957:Plasmids; D017433:Protein Structure, Secondary; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D017386:Sequence Homology, Amino Acid; D015534:Trans-Activators / Q000276:immunology / Q000378:metabolism*; D014157:Transcription Factors; D016335:Zinc Fingers / Q000235:genetics
D000906:Antibodies; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C052130:GAL4 protein, S cerevisiae; C058452:HAP1 protein, S cerevisiae; D046911:Macromolecular Substances; D029701:Saccharomyces cerevisiae Proteins; D015534:Trans-Activators; D014157:Transcription Factors; D006418:Heme