The wing in yeast heat shock transcription factor (HSF) DNA-binding domain is required for full activity

The yeast heat shock transcription factor (HSF) belongs to the winged helix family of proteins. HSF binds DNA as a trimer, and additional trimers can bind DNA co-operatively. Unlike other winged helix–turn–helix proteins, HSF’s wing does not appear to contact DNA, as based on a previously solved crystal structure. Instead, the structure implies that the wing is involved in protein–protein interactions, possibly within a trimer or between adjacent trimers. To understand the function of the wing in the HSF DNA-binding domain, a Saccharomyces cerevisiae strain was created that expresses a wingless HSF protein. This strain grows normally at 30°C, but shows a decrease in reporter gene expression during constitutive and heat-shocked conditions. Removal of the wing does not affect the stability or trimeric nature of a protein fragment containing the DNA-binding and trimerization domains. Removal of the wing does result in a decrease in DNA-binding affinity. This defect was mainly observed in the ability to form the first trimer-bound complex, as the formation of larger complexes is unaffected by the deletion. Our results suggest that the wing is not involved in the highly co-operative nature of HSF binding, but may be important in stabilizing the first trimer bound to DNA.

酵母热休克转录因子（HSF）隶属于翼螺旋蛋白家族（winged helix family）。HSF以三聚体形式结合DNA，且额外的三聚体可协同结合DNA。与其他翼螺旋-转角-螺旋（winged helix-turn-helix）蛋白不同，根据已解析的晶体结构，HSF的翼状结构域似乎并不与DNA接触。反之，该结构提示该翼状结构域参与蛋白质-蛋白质相互作用，可能发生在三聚体内部或相邻三聚体之间。为探究HSF DNA结合结构域中翼状结构域的功能，研究人员构建了一株表达无翼状结构域HSF蛋白的酿酒酵母（Saccharomyces cerevisiae）菌株。该菌株在30℃下可正常生长，但在组成型表达及热激条件下，报告基因的表达量均出现下降。移除该翼状结构域并不会影响包含DNA结合结构域与三聚化结构域的蛋白质片段的稳定性及三聚化特性，但确实会降低其DNA结合亲和力。该缺陷主要体现在首个三聚体结合复合物的形成能力上，而更大复合物的形成过程则不受该缺失突变的影响。本研究结果表明，翼状结构域并不参与HSF结合DNA的高协同性过程，但可能对稳定首个结合至DNA的三聚体具有重要作用。