Department of Chemistry, Washington University, St. Louis, Missouri, United States of America.
PLoS One. 2020 Mar 10;15(3):e0230198. doi: 10.1371/journal.pone.0230198. eCollection 2020.
Hsp104 is a hexameric AAA+ yeast disaggregase capable of solubilizing disordered aggregates and amyloid. Hsp104 couples ATP hydrolysis to polypeptide translocation through its central channel. Substrate binding by Hsp104 is mediated primarily by two conserved tyrosine residues in nucleotide binding domain (NBD) 1 and NBD2. Recent structural studies have revealed that an additional tyrosine residue (Y650) located in NBD2 appears to contact substrate and may play an important role in Hsp104 function. Here, we functionally analyze the properties of this proposed Hsp104 -substrate interaction. We find that Y650 is not essential for Hsp104 to confer thermotolerance. Supporting these findings, in a potentiated Hsp104 variant background, the Y650A mutation does not abolish potentiation. However, modulation of this site does have subtle effects on the activity of this potentiated Hsp104 variant. We therefore suggest that while Y650 is not essential for Hsp104 function, its modulation may be useful for fine-tuning Hsp104 properties.
Hsp104 是一种六聚体的 AAA+ 酵母解聚酶，能够溶解无规聚集物和淀粉样蛋白。Hsp104 通过其中心通道将 ATP 水解与多肽易位偶联。Hsp104 的底物结合主要由核苷酸结合域 (NBD) 1 和 NBD2 中的两个保守的酪氨酸残基介导。最近的结构研究表明，位于 NBD2 中的另一个酪氨酸残基 (Y650) 似乎与底物接触，并可能在 Hsp104 功能中发挥重要作用。在这里，我们从功能上分析了这个拟议的 Hsp104-底物相互作用的特性。我们发现 Y650 对于 Hsp104 赋予耐热性不是必需的。支持这些发现，在增强型 Hsp104 变体背景下，Y650A 突变并没有消除增强作用。然而，该位点的调节对这种增强型 Hsp104 变体的活性有细微的影响。因此，我们认为虽然 Y650 对于 Hsp104 功能不是必需的，但它的调节可能有助于微调 Hsp104 的特性。
