Department of Nanotechnology, Islamic Azad University of Pharmaceutical Sciences Branch, Tehran, Iran.
Department of clinical Biochemistry and Genetics, School of Medicine, Qazvin University of Medical Sciences, Qazvin, Iran.
Biochim Biophys Acta Proteins Proteom. 2017 Mar;1865(3):370-379. doi: 10.1016/j.bbapap.2016.12.013. Epub 2016 Dec 23.
Tyrosinase is a determinant enzyme for modulating melanin production as its abnormal activity can result in an increased amount of melanin. Reduction of tyrosinase activity has been targeted for preventing and healing hyperpigmentation of skin, such as melanoma and age related spots. The aim of this systematic study is to investigate whether recombinant S100A8/A9 and its modified form reduce the activity of mushroom tyrosinase (MT) through changing its structure. Recombinant His-Tagged S100A8 and S100A9 are expressed in Escherichia coli BL21 (DE3) and modified using Woodward's reagent K which is a carboxyl group modifier. The structures of S100A8/A9 and its modified form are studied using fluorescence and circular dichroism spectroscopy, and the activity of MT is measured using UV-visible spectrophotometry in the presence of its substrate, L-3,4-dihydroxyphenylalanine (L-DOPA). The results show a lower stability of the modified protein when compared with its unmodified form. The interaction of S100A8/A9 with MT changes the structure and successfully reduces the activity of mushroom tyrosinase. Recombinant S100A8/A9 complex decreases MT activity which can control malignant melanoma, the most dangerous type of skin cancer.
酪氨酸酶是调节黑色素生成的决定性酶，因为其异常活性会导致黑色素生成增加。抑制酪氨酸酶的活性已被用于预防和治疗皮肤色素沉着过度，如黑色素瘤和与年龄相关的斑点。本系统研究的目的是研究重组 S100A8/A9 及其修饰形式是否通过改变结构来降低蘑菇酪氨酸酶（MT）的活性。重组 His 标记的 S100A8 和 S100A9 在大肠杆菌 BL21（DE3）中表达，并使用 Woodward 试剂 K 进行修饰，Woodward 试剂 K 是一种羧基修饰剂。使用荧光和圆二色性光谱研究 S100A8/A9 及其修饰形式的结构，并在其底物 L-3,4-二羟基苯丙氨酸（L-DOPA）存在下使用紫外可见分光光度法测量 MT 的活性。结果表明，与未修饰形式相比，修饰蛋白的稳定性较低。S100A8/A9 与 MT 的相互作用改变了结构，并成功降低了蘑菇酪氨酸酶的活性。重组 S100A8/A9 复合物降低 MT 活性，可控制恶性黑色素瘤，这是最危险的皮肤癌类型。
