Altering the DNA-binding specificity of the yeast Matalpha 2 homeodomain protein.
Homeodomain proteins are a highly conserved class of DNA-binding proteins that are found in virtually every eukaryotic organism. The conserved mechanism that these proteins use to bind DNA suggests that there may be at least a partial DNA recognition code for this class of proteins. To test this idea, we have investigated the sequence-specific requirements for DNA binding and repression by the yeast alpha2 homeodomain protein in association with its cofactors, Mcm1 and Mata1. We have determined the contribution for each residue in the alpha2 homeodomain that contacts the DNA in the co-crystal structures of the protein. We have also engineered mutants in the alpha2 homeodomain to alter the DNA-binding specificity of the protein. Although we were unable to change the specificity of alpha2 by making substitutions at residues 47, 54, and 55, we were able to alter the DNA-binding specificity by making substitutions at residue 50 in the homeodomain. Since other homeodomain proteins show similar changes in specificity with substitutions at residue 50, this suggests that there is at least a partial DNA recognition code at this position.
Mathias
J R
JR
Waksman Institute and the Department of Molecular Biology and Biochemistry, Rutgers University, Piscataway, New Jersey 08854-8020, USA.
Zhong
H
H
Jin
Y
Y
Vershon
A K
AK
D019943:Amino Acid Substitution; D001665:Binding Sites; D018360:Crystallography, X-Ray; D015342:DNA Probes; D004271:DNA, Fungal / Q000737:chemistry* / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D018398:Homeodomain Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D026341:Minichromosome Maintenance 1 Protein; D016297:Mutagenesis, Site-Directed; D009690:Nucleic Acid Conformation; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012097:Repressor Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D012694:Serine; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D015342:DNA Probes; D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D018398:Homeodomain Proteins; C096063:MATA1 protein, S cerevisiae; C062928:MATA2 protein, S cerevisiae; D026341:Minichromosome Maintenance 1 Protein; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D012694:Serine; D001616:beta-Galactosidase