TY - JOUR
T1 - The TATA-binding protein from Saccharomyces cerevisiae oligomerizes in solution at micromolar concentrations to form tetramers and octamers
AU - Daugherty, Margaret A.
AU - Brenowitz, Michael
AU - Fried, Michael G.
N1 - Funding Information:
Supported by NSF Grant DMB 91-48816 to M.G.F., a Four Diamonds Fund Postdoctoral Grant to M.A.D. and NIH GM51506 and GM39929 to M.B.
PY - 1999/1/29
Y1 - 1999/1/29
N2 - Equilibrium analytical ultracentrifugation has been used to determine the stoichiometry and energetics of the self-assembly of the TATA-binding protein of Saccharomyces cerevisiae at 30°C, in buffers ranging in salt concentration from 60 mM KCl to 1 M KCl. The data are consistent with a sequential association model in which monomers are in equilibrium with tetramers and octamers at protein concentrations above 2.6 μM. Association is highly cooperative, with octamer formation favored by ~7 kcal/mol over tetramers. At high [KCl], the concentration of tetramers becomes negligible and the data are best described by a monomer-octamer reaction mechanism. The equilibrium association constants for both monomer⇆tetramer and tetramer⇆octamer reactions change with [KCl] in a biphasic manner, decreasing with increasing [KCl] from 60 mM to 300 mM, and increasing with increasing [KCl] from 300 mM to 1 M. At low [KCl], ~3 mole equivalents of ions are released at each association step, while at high [KCl], ~3 mole equivalents of ions are taken up at each association step. These results suggest that there is a salt concentration-dependent change in the assembly mechanism, and that the mechanistic switch takes place near 300 mM KCl. The possibility that this self-association reaction may play a role in the activity of the TATA-binding protein in vivo is discussed.
AB - Equilibrium analytical ultracentrifugation has been used to determine the stoichiometry and energetics of the self-assembly of the TATA-binding protein of Saccharomyces cerevisiae at 30°C, in buffers ranging in salt concentration from 60 mM KCl to 1 M KCl. The data are consistent with a sequential association model in which monomers are in equilibrium with tetramers and octamers at protein concentrations above 2.6 μM. Association is highly cooperative, with octamer formation favored by ~7 kcal/mol over tetramers. At high [KCl], the concentration of tetramers becomes negligible and the data are best described by a monomer-octamer reaction mechanism. The equilibrium association constants for both monomer⇆tetramer and tetramer⇆octamer reactions change with [KCl] in a biphasic manner, decreasing with increasing [KCl] from 60 mM to 300 mM, and increasing with increasing [KCl] from 300 mM to 1 M. At low [KCl], ~3 mole equivalents of ions are released at each association step, while at high [KCl], ~3 mole equivalents of ions are taken up at each association step. These results suggest that there is a salt concentration-dependent change in the assembly mechanism, and that the mechanistic switch takes place near 300 mM KCl. The possibility that this self-association reaction may play a role in the activity of the TATA-binding protein in vivo is discussed.
KW - Self-association
KW - TBP
KW - Thermodynamics
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U2 - 10.1006/jmbi.1998.2427
DO - 10.1006/jmbi.1998.2427
M3 - Article
C2 - 9917384
AN - SCOPUS:0033613911
SN - 0022-2836
VL - 285
SP - 1389
EP - 1399
JO - Journal of Molecular Biology
JF - Journal of Molecular Biology
IS - 4
ER -