Direct interaction of tyrosinase with Tyrp1 to form heterodimeric complexes in vivo
Direct interaction of tyrosinase with Tyrp1 to form heterodimeric complexes in vivo
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DOI：
10.1242/jcs.017913
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发表时间：
2007-12-15
影响因子：
4
通讯作者：
Hearing, Vincent J.
中科院分区：
文献类型：
--
作者：
Kobayashi, Takeshi;Hearing, Vincent J.
关键的和限速的黑素生成酶酪氨酸酶（Tyr）的突变导致头发、皮肤和眼睛的色素减退。另外两种相关的酶，Tyrp 1和Dct，催化黑色素生物合成中不同的后-Tyr反应。已经提出Tyr、Tyrp 1和Dct在多酶复合物中相互作用并相互稳定，并且在体外，Tyr活性在Tyrp 1和/或Dct存在下更稳定。我们最近报道，Tyr在突变型Tyrp 1小鼠黑素细胞中比在野生型Tyrp 1黑素细胞中降解得更快，并且Tyr的稳定性降低可以通过感染野生型Tyrp 1而部分挽救。虽然Tyr和Tyrp 1之间的相互作用已在体外得到证实，但没有直接证据表明Tyr与Tyrp 1在体内相互作用。在这项研究中，我们使用体内化学交联来稳定Tyr与其他细胞蛋白的结合。蛋白质印迹分析表明，Tyrp 1，但不是Dct，与酪氨酸在小鼠黑素细胞在体内，更具体地说，在黑素体。二维SDS-PAGE分析检测到Tyr和Tyrp 1的异二聚体种类。总之，这些数据表明，Tyrp 1直接相互作用与酪氨酸在体内，这可能会调节稳定性和运输的黑素生成酶，从而色素合成。
Mutations of the critical and rate-limiting melanogenic enzyme tyrosinase (Tyr) result in hypopigmentation of the hair, skin and eyes. Two other related enzymes, Tyrp1 and Dct, catalyze distinct post-Tyr reactions in melanin biosynthesis. Tyr, Tyrp1 and Dct have been proposed to interact with and stabilize each other in multi-enzyme complexes, and in vitro, Tyr activity is more stable in the presence of Tyrp1 and/or Dct. We recently reported that Tyr is degraded more quickly in mutant Tyrp1 mouse melanocytes than in wild-type Tyrp1 melanocytes, and that decreased stability of Tyr can be partly rescued by infection with wild-type Tyrp1. Although interactions between Tyr and Tyrp1 have been demonstrated in vitro, there is no direct evidence for Tyr interaction with Tyrp1 in vivo. In this study, we use in vivo chemical crosslinking to stabilize the association of Tyr with other cellular proteins. Western blot analysis revealed that Tyrp1, but not Dct, associates with Tyr in murine melanocytes in vivo, and more specifically, in melanosomes. Two-dimensional SDS-PAGE analysis detected heterodimeric species of Tyr and Tyrp1. Taken together, these data demonstrate that Tyrp1 interacts directly with Tyr in vivo, which may regulate the stability and trafficking of melanogenic enzymes and thus pigment synthesis.