SELECTIVE PROTEOLYSIS OF ARRESTIN BY CALPAIN - MOLECULAR CHARACTERISTICS AND ITS EFFECT ON RHODOPSIN DEPHOSPHORYLATION
SELECTIVE PROTEOLYSIS OF ARRESTIN BY CALPAIN - MOLECULAR CHARACTERISTICS AND ITS EFFECT ON RHODOPSIN DEPHOSPHORYLATION
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DOI：
10.1074/jbc.270.41.24375
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发表时间：
1995-10-13
影响因子：
4.8
通讯作者：
WILLIAMS, DS
中科院分区：
文献类型：
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作者：
AZARIAN, SM;KING, AJ;WILLIAMS, DS
视觉抑制蛋白（48 kDa）通过与光激活的磷酸化形式的受体结合，在视紫红质的失活中发挥作用。在蛋白酶抑制剂存在下制备的牛视杆外节中，通过Western印迹分析观察到两种更快迁移形式的抑制蛋白，其表观分子量为46和44 kDa。46-kDa形式在保持在光照下的眼的视杆外节中比放置在光照下的眼的视杆外节中更明显，在黑暗中，发现与通过纯视网膜钙蛋白酶II体外蛋白水解arrestin产生的相同。体外分析表明，arrestin蛋白水解时，只有结合到视紫红质;可溶性arrestin没有显着切割钙蛋白酶。蛋白水解涉及在两个，可能三个位点的连续切割，导致从COOH末端去除27个氨基酸。剩余的46-kDa蛋白质抵抗钙蛋白酶的进一步蛋白水解。与完整的arrestin不同，46-kDa截短的arrestin在受体失去其发色团后不容易从受体释放，也不容易在加入11-顺式-视黄醛以再生受体后释放。发现截短的arrestin抑制受体去磷酸化的程度与完整的arrestin相同。总之，这些结果提供的证据表明，一个46 kDa形式的抑制蛋白在杆外节是一个产品的选择性蛋白水解钙蛋白酶。此外，他们认为这种蛋白水解可能提供了一种延长视觉受体磷酸化状态的机制。
Visual arrestin (48 kDa) plays a role in the deactivation of rhodopsin by binding to the light-activated, phosphorylated form of the receptor. In bovine rod outer segments that were prepared in the presence of protease inhibitors, two faster migrating forms of arrestin, with apparent molecular masses of 46 and 44 kDa, were observed by Western blot analysis, The 46-kDa form was more evident in rod outer segments of eyes kept in the light than those placed, in darkness and was found to be identical to that generated by in vitro proteolysis of arrestin by pure retinal calpain II. In vitro analysis showed that arrestin was proteolyzed only when bound to rhodopsin; soluble arrestin was not significantly cleaved by calpain. Proteolysis involves sequential cleavage at two, possibly three sites, resulting in the removal of 27 amino acids from the COOH terminus. The remaining 46-kDa protein was resistant to further proteolysis by calpain. Unlike intact arrestin, the 46-kDa truncated arrestin was not readily released from the receptor after the receptor had lost its chromophore, nor was it released upon the addition of 11-cis-retinal to regenerate the receptor. Truncated arrestin was found to inhibit receptor dephosphorylation to the same extent as intact arrestin. In conclusion, these results provide evidence that a 46 kDa form of arrestin in rod outer segments is a product of selective proteolysis by calpain. Furthermore, they suggest that this proteolysis may provide a mechanism for prolonging the phosphorylated state of the visual receptor.