Calcium, nucleotide, and actin affect the interaction of mammalian Myo1c with its light chain calmodulin.
Calcium, nucleotide, and actin affect the interaction of mammalian Myo1c with its light chain calmodulin.
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钙、核苷酸和肌动蛋白影响哺乳动物 Myo1c 与其轻链钙调蛋白的相互作用。
DOI：
10.1021/bi8011059
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发表时间：
2008
期刊：
影响因子：
2.9
通讯作者：
Coluccio,LynneM
中科院分区：
文献类型：
--
作者：
Lieto-Trivedi,Alena;Coluccio,LynneM
为了研究哺乳动物I类肌球蛋白Myo1c与其轻链钙调蛋白的相互作用，我们表达了由Myo1c运动结构域和0−4个假定的钙调蛋白结合（IQ）结构域（Myo1c0IQ−Myo1c4IQ）组成的截断突变体。从myo1c0iq到Myo1c3IQ，随着IQ结构域数量的增加，与Myo1c重链相关的钙调蛋白的数量也随之增加。除了与myo1c3iq相关的钙调蛋白外，没有发现与myo1c4iq相关的钙调蛋白，尽管它是可用的，这表明Myo1c结合了三个钙调蛋白分子，没有证据表明存在第四个智商结构域。与Myo1c0IQ不同的是，myo1c1iq的基础atp酶活性在Ca2+条件下比EGTA±外源性钙调蛋白高10倍，表明Ca2+结合钙调蛋白在第一个IQ结构域进行调控。肌动蛋白激活的Mg2+- atp酶活性的kmandvmax在很大程度上独立于存在的IQ结构域的数量，并受到Ca2+的适度影响。在结合实验中，当肌动蛋白存在时，一些钙调蛋白与Myo1c重链形成颗粒，但相当一部分仍留在上清中，这表明钙调蛋白最有可能从第二IQ结构域转移。Myo1c重链以核苷酸依赖的方式与肌动蛋白相关。在ATP中，较小比例的钙调蛋白与重链形成颗粒，这表明Myo1c经历了核苷酸依赖的构象变化，影响了钙调蛋白对重链的亲和力。这些研究支持一种模型，即内耳的Myo1c受Ca2+和核苷酸的调节，它们通过轻链结合区对运动活动产生影响。
To investigate the interaction of mammalian class I myosin, Myo1c, with its light chain calmodulin, we expressed (with calmodulin) truncation mutants consisting of the Myo1c motor domain followed by 0−4 presumed calmodulin-binding (IQ) domains (Myo1c0IQ−Myo1c4IQ). The amount of calmodulin associating with the Myo1c heavy chain increased with increasing number of IQ domains from Myo1c0IQto Myo1c3IQ. No calmodulin beyond that associated with Myo1c3IQwas found with Myo1c4IQdespite its availability, showing that Myo1c binds three molecules of calmodulin with no evidence of a fourth IQ domain. Unlike Myo1c0IQ, the basal ATPase activity of Myo1c1IQwas >10-fold higher in Ca2+vs EGTA ± exogenous calmodulin, showing that regulation is by Ca2+binding to calmodulin on the first IQ domain. TheKmandVmaxof the actin-activated Mg2+-ATPase activity were largely independent of the number of IQ domains present and moderately affected by Ca2+. In binding assays, some calmodulin pelleted with Myo1c heavy chain when actin was present, but a considerable fraction remained in the supernatant, suggesting that calmodulin is displaced most likely from the second IQ domain. The Myo1c heavy chain associated with actin in a nucleotide-dependent fashion. In ATP a smaller proportion of calmodulin pelleted with the heavy chain, suggesting that Myo1c undergoes nucleotide-dependent conformational changes that affect the affinity of calmodulin for the heavy chain. The studies support a model in which Myo1c in the inner ear is regulated by both Ca2+and nucleotide, which exert their effects on motor activity through the light-chain-binding region.