Self-association of the amino-terminal domain of the yeast TATA-binding protein.
The amino-terminal domain of yeast TATA-binding protein has been proposed to play a crucial role in the self-association mechanism(s) of the full-length protein. Here we tested the ability of this domain to self-associate under a variety of solution conditions. Escherichia coli two-hybrid assays, in vitro pull-down assays, and in vitro cross-linking provided qualitative evidence for a limited and specific self-association. Sedimentation equilibrium analysis using purified protein was consistent with a monomer-dimer equilibrium with an apparent dissociation constant of approximately 8.4 microM. Higher stoichiometry associations remain possible but could not be detected by any of these methods. These results demonstrate that the minimal structure necessary for amino-terminal domain self-association must be present even in the absence of carboxyl-terminal domain structures. On the basis of these results we propose that amino-terminal domain structures contribute to the oligomerization interface of the full-length yeast TATA-binding protein.
Adams
Claire A
CA
Department of Biochemistry and Molecular Biology, The Pennsylvania State University College of Medicine, Hershey, Pennsylvania 17033, USA.
Kar
Sambit R
SR
Hopper
James E
JE
Fried
Michael G
MG
D003001:Cloning, Molecular; D003432:Cross-Linking Reagents / Q000494:pharmacology; D017931:DNA Primers / Q000737:chemistry; D012321:DNA-Directed RNA Polymerases / Q000737:chemistry; D019281:Dimerization; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000378:metabolism; D005656:Fungal Proteins / Q000737:chemistry; D005822:Genetic Vectors; D007700:Kinetics; D008957:Models, Genetic; D015233:Models, Statistical; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D035181:TATA-Box Binding Protein / Q000737:chemistry*; D020798:Two-Hybrid System Techniques
D003432:Cross-Linking Reagents; D017931:DNA Primers; D005656:Fungal Proteins; D035181:TATA-Box Binding Protein; D012321:DNA-Directed RNA Polymerases