Department of Neuropathology and Neuroscience, Graduate School of Pharmaceutical Sciences, The University of Tokyo , 7-3-1 Hongo, Bunkyoku, Tokyo, 113-0033 Japan.
Biochemistry. 2013 Sep 3;52(35):6052-62. doi: 10.1021/bi400596m. Epub 2013 Aug 23.
Mutations in the leucine-rich repeat kinase 2 (LRRK2) gene have been identified in pedigrees of autosomal-dominant familial Parkinson's disease (PARK8). It has been shown that the kinase activity of LRRK2 is required for its neuronal toxicity, although how familial Parkinson mutations affect the function of LRRK2 has not been well characterized. In the present study, we systematically characterized the autophosphorylation of LRRK2 by phosphopeptide mapping and identified Thr1348, Thr1349, and Thr1357 as the major autophosphorylation sites. We found that the autophosphorylation at Thr1357 is downregulated by the Y1699C mutation, possibly through a conformational alteration of the ROC domain. We also found that I2020T mutant LRRK2 undergoes excessive autophosphorylation in cell lysates in vitro at a low concentration of ATP. These results highlight the differential effects of familial mutations in LRRK2 on its conformation and enzymatic properties.
LRRK2 基因中的突变已在常染色体显性家族性帕金森病（PARK8）的家系中被鉴定出来。已经表明，LRRK2 的激酶活性对于其神经元毒性是必需的，尽管家族性帕金森突变如何影响 LRRK2 的功能尚未得到很好的描述。在本研究中，我们通过磷酸肽图谱系统地描述了 LRRK2 的自身磷酸化，并鉴定出 Thr1348、Thr1349 和 Thr1357 是主要的自身磷酸化位点。我们发现，Y1699C 突变使 Thr1357 的自身磷酸化下调，可能是通过 ROC 结构域的构象改变。我们还发现，I2020T 突变的 LRRK2 在体外低浓度 ATP 时在细胞裂解物中发生过度的自身磷酸化。这些结果突出了家族性 LRRK2 突变对其构象和酶特性的不同影响。
