The hydroxyl group of S685 in Walker A motif and the carboxyl group of D792 in Walker B motif of NBD1 play a crucial role for multidrug resistance protein folding and function.
The hydroxyl group of S685 in Walker A motif and the carboxyl group of D792 in Walker B motif of NBD1 play a crucial role for multidrug resistance protein folding and function.
复制标题
NBD1的Walker A基序中S685的羟基和Walker B基序中D792的羧基对于多药耐药蛋白的折叠和功能起着至关重要的作用。
DOI：
10.1016/j.bbamem.2007.11.010
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发表时间：
2008
期刊：
影响因子：
--
通讯作者：
Chang,Xiu-Bao
中科院分区：
文献类型：
--
作者：
Yang,Runying;Scavetta,Robert;Chang,Xiu-Bao
MRP1-NBD1的结构分析表明，Walker A S685与Walker B D792形成氢键，并与镁和结合ATP的β-磷酸相互作用。我们发现用亮氨酸取代D792会导致蛋白质的错误折叠。在本报告中，我们测试了用阻止氢键形成的残基取代S685是否也会导致错误折叠。事实上，用可能阻止氢键形成的残基取代S685会导致蛋白质的错误折叠。此外，一些可能与D792形成氢键的取代也产生了未成熟蛋白。所有这些变种都是对温度敏感的变种。然而，从27°C培养的细胞中制备的这些复杂糖基化的成熟突变体仍然显著影响ATP结合和ATP依赖的溶质运输。相比之下，用苏氨酸取代S685产生了比野生型MRP1更活跃的复杂糖基化成熟蛋白，这表明685残基羟基与D792羧基之间的相互作用对蛋白质折叠起着至关重要的作用，685羟基与镁和结合ATP的β-磷酸的相互作用对ATP结合和ATP依赖的溶质转运起着重要作用。
Structural analysis of MRP1-NBD1 revealed that the Walker A S685 forms hydrogen-bond with the Walker B D792 and interacts with magnesium and the β-phosphate of the bound ATP. We have found that substitution of the D792 with leucine resulted in misfolding of the protein. In this report we tested whether substitution of the S685 with residues that prevent formation of this hydrogen-bond would also cause misfolding. Indeed, substitution of the S685 with residues potentially preventing formation of this hydrogen-bond resulted in misfolding of the protein. In addition, some substitutions that might form hydrogen-bond with D792 also yielded immature protein. All these mutants are temperature-sensitive variants. However, these complex-glycosylated mature mutants prepared from the cells grown at 27 °C still significantly affect ATP binding and ATP-dependent solute transport. In contrast, substitution of the S685 with threonine yielded complex-glycosylated mature protein that is more active than the wild-type MRP1, indicating that the interaction between the hydroxyl group of 685 residue and the carboxyl group of D792 plays a crucial role for the protein folding and the interactions of the hydroxyl group at 685 with magnesium and the β-phosphate of the bound ATP play an important role for ATP-binding and ATP-dependent solute transport.