Zerovnik Eva, Skerget Katja, Tusek-Znidaric Magda, Loeschner Corina, Brazier Marcus W, Brown David R
Department of Biochemistry and Molecular Biology, JoZef Stefan Institute, Ljubljana, Slovenia.
FEBS J. 2006 Sep;273(18):4250-63. doi: 10.1111/j.1742-4658.2006.05426.x.
We show that human stefin B, a protease inhibitor from the family of cystatins, is a copper binding protein, unlike stefin A. We have used isothermal titration calorimetry to directly monitor the binding event at pH 7 and pH 5. At pH 7 stefin B shows a picomolar affinity for copper but at pH 5 the affinity is in the nanomolar range. There is no difference in the affinity of copper between the wildtype stefin B (E31 isoform) and a variant (Y31 isoform), whereas the mutant (P79S), which is tetrameric, does not bind copper. The conformation of stefin B remains unaltered by copper binding. It is known that below pH 5 stefin B undergoes a conformational change and amyloid fibril formation. We show that copper binding inhibits the amyloid fibril formation and, to a lesser degree, the initial aggregation. Similarities to and differences from other copper binding amyloidogenic proteins are discussed.
我们发现，与丝抑蛋白A不同，人丝抑蛋白B（一种来自半胱氨酸蛋白酶抑制剂家族的蛋白酶抑制剂）是一种铜结合蛋白。我们使用等温滴定量热法直接监测了在pH 7和pH 5时的结合事件。在pH 7时，丝抑蛋白B对铜表现出皮摩尔亲和力，但在pH 5时，亲和力处于纳摩尔范围。野生型丝抑蛋白B（E31异构体）和变体（Y31异构体）对铜的亲和力没有差异，而四聚体的突变体（P79S）不结合铜。丝抑蛋白B的构象不会因铜结合而改变。已知在pH 5以下，丝抑蛋白B会发生构象变化并形成淀粉样纤维。我们发现铜结合会抑制淀粉样纤维的形成，并在较小程度上抑制初始聚集。文中还讨论了与其他铜结合淀粉样蛋白生成蛋白的异同。
