大约99%的线粒体蛋白质组是细胞核编码的,在细胞质中合成,随后导入并分类到细胞器的正确隔室中。线粒体内膜23(TIM23)复合物的转位酶是内膜的主要蛋白质转位酶,并负责蛋白质穿过内膜并插入内膜。Tim23是构成导入通道的综合体的中心组件。仍然缺少进口通道的高分辨率结构,对其功能重要的结构元素是未知的。在本研究中,我们分析了Tim23跨膜片段中高度丰富的GxxxG基序对于TIM23复合物结构完整性的重要性.在第一个GxxxG基序中存在的10个甘氨酸中,Tim23的第二和第三跨膜片段,其中三个在跨膜片段1和2中的突变导致致死表型,和温度敏感表型中的其他三个突变。其余四种无明显生长表型。重要的是,这些突变均未损害Tim23前体向线粒体的导入和膜整合。然而,生长障碍的严重程度与TIM23复合体的不稳定相关。我们得出的结论是,在Tim23的第一和第二跨膜片段中发现的GxxxG基序对于TIM23复合物的结构完整性是必需的。
Approximately 99% of the mitochondrial proteome is nucleus-encoded, synthesized in the cytosol, and subsequently imported into and sorted to the correct compartment in the organelle. The translocase of the inner mitochondrial membrane 23 (TIM23) complex is the major protein translocase of the inner membrane, and is responsible for translocation of proteins across the inner membrane and their insertion into the inner membrane. Tim23 is the central component of the complex that forms the import channel. A high-resolution structure of the import channel is still missing, and structural elements important for its function are unknown. In the present study, we analyzed the importance of the highly abundant GxxxG motifs in the transmembrane segments of Tim23 for the structural integrity of the TIM23 complex. Of 10 glycines present in the GxxxG motifs in the first, second and third transmembrane segments of Tim23, mutations of three of them in transmembrane segments 1 and 2 resulted in a lethal phenotype, and mutations of three others in a temperature-sensitive phenotype. The remaining four caused no obvious growth phenotype. Importantly, none of the mutations impaired the import and membrane integration of Tim23 precursor into mitochondria. However, the severity of growth impairment correlated with the destabilization of the TIM23 complex. We conclude that the GxxxG motifs found in the first and second transmembrane segments of Tim23 are necessary for the structural integrity of the TIM23 complex.