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首页> 外文期刊>Biochemistry >Dimer dissociation and thermosensitivity kinetics of the Saccharomyces cerevisiae and human TATA binding proteins.
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Dimer dissociation and thermosensitivity kinetics of the Saccharomyces cerevisiae and human TATA binding proteins.

机译：酿酒酵母和人TATA结合蛋白的二聚体解离和热敏动力学。

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A kinetic analysis of dimer dissociation, TATA DNA binding, and thermal inactivation of the yeast Saccharomyces cerevisiae and human TATA binding proteins (TBP) was conducted. We find that yeast TBP dimers, like human TBP dimers, are slow to dissociate in vitro (t(1/2) approximately 20 min). Mild mutations in the crystallographic dimer interface accelerate the rate of dimer dissociation, whereas severe mutations prevent dimerization. In the presence of excess TATA DNA, which measures the entire active TBP population, dimer dissociation represents the rate-limiting step in DNA binding. These findings provide a biochemical extension to genetic studies demonstrating that TBP dimerization prevents unregulated gene expression in yeast [Jackson-Fisher, A. J., Chitikila, C., Mitra, M., and Pugh, B. F. (1999) Mol. Cell 3, 717-727]. In the presence of vast excesses of TBP over TATA DNA, which measures only a very small fraction of the total TBP, the monomer population in a monomer/dimer equilibrium binds DNA rapidly, which is consistent with a simultaneous binding and bending of the DNA. Under conditions where other studies failed to detect dimers, yeast TBP's DNA binding activity was extremely labile in the absence of TATA DNA, even at temperatures as low as 0 degrees C. Kinetic analyses of TBP instability in the absence of DNA at 30 degrees C revealed that even under fairly stabilizing solution conditions, TBP's DNA binding activity rapidly dissipated with t(1/2) values ranging from 6 to 26 min. TBP's stability appeared to vary with the square root of the TBP concentration, suggesting that TBP dimerization helps prevent TBP inactivation.
机译：对酵母酿酒酵母和人TATA结合蛋白（TBP）进行了二聚体解离，TATA DNA结合和热灭活的动力学分析。我们发现酵母TBP二聚体，像人类TBP二聚体，是缓慢离解离体（t（1/2）大约20分钟）。晶体学二聚体界面中的轻度突变会加速二聚体的解离速率，而重度突变则会阻止二聚体化。在测量整个活性TBP群体的过量TATA DNA存在下，二聚体解离代表DNA结合中的限速步骤。这些发现提供了遗传学研究的生化扩展，证明了TBP二聚化阻止了酵母中不受调节的基因表达[Jackson-Fisher，A.J.，Chitikila，C.，Mitra，M。，和Pugh，B.F。（1999）Mol.Biol.215：403-10。单元3，717-727]。在TBP相对于TATA DNA而言仅存在非常少量的TBP的情况下，TBA仅占总TBP的很小一部分，单体/二聚体平衡中的单体群体会迅速结合DNA，这与DNA的同时结合和弯曲一致。在其他研究未能检测到二聚体的条件下，即使在低至0摄氏度的温度下，即使没有TATA DNA，酵母TBP的DNA结合活性也极不稳定。在30摄氏度下不存在DNA的情况下，TBP不稳定的动力学分析表明即使在相当稳定的溶液条件下，TBP的DNA结合活性也迅速消失，t（1/2）值在6至26分钟之间。 TBP的稳定性似乎随TBP浓度的平方根而变化，这表明TBP二聚化有助于防止TBP失活。

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