Molecular design of the single-headed processive myosin

马达蛋白根据其结构和行为属于以下类别之一：双头非进行性，双头进行性，单头非进行性和单头进行性。肌球蛋白II是第一类。肌球蛋白V和VI属于第二类。肌球蛋白I属于第三类。驱动蛋白超家族中的一种蛋白属于最后一类。在人类肌球蛋白中，肌球蛋白IIIA、肌球蛋白IIIB、肌球蛋白IXA和肌球蛋白IXA被认为属于最后一类，尽管尚未通过实验确定这些肌球蛋白是否真的是单头和进行性的。我们从人脑cDNA文库中克隆了人肌球蛋白IIIA、IIIB和IXB的全长cDNA。克隆的肌球蛋白IIIA和IXB cDNA表明存在几种类型的选择性剪接。我们已经在Sf9或high five细胞中表达了这些cDNA，包括几种剪接变体。从转染的细胞中，我们已经纯化了表达的蛋白质用于进一步的生化和生物物理分析。在这个阶段，由于包涵体的形成，我们只能部分成功地分离蛋白质。已经尝试了各种纯化方法来增加表达的蛋白质的溶解度。需要进一步的研究，以最终确定肌球蛋白IIIA，IIIB和IXB的运动特性。

Motor proteins belong one of the classes according to their structure and behavior ; two-headed non-processive, two-headed processive, one-headed non-processive and one-headed processive. Myosin II is in the first class. Myosin V and VI belong to the second class. Myosin I belongs to the third class. One of the kinesin superfamily proteins belong to the last class. In human myosins, myosin IIIA, myosin IIIB, myosin IXA, and myosin IXA are supposed to belong to the last class, although it has not yet been determined experimentally if these myosins are actually one-headed and processive. We have cloned full-length cDNA of human myosin IIIA, IIIB, and IXB from the human brain cDNA library. Cloned myosin IIIA and IXB cDNAs show that there are several types of alternative splicing. We have expressed these cDNAs including several splice variants in Sf9 or high five cells. From the transfected cells, we have purified the expressed proteins for further biochemical and biophysical analyzes. At this stage, we are only partially successful to isolate the proteins due to the formation of inclusion bodies. Various purification procedures have been tried to increase the solubility of expressed proteins. Further studies are required to finally identify the motor characteristics of myosin IIIA, IIIB and IXB.