Structural and Functional Dissection of Mif2p, a Conserved DNA-binding Kinetochore Protein
Structural and Functional Dissection of Mif2p, a Conserved DNA-binding Kinetochore Protein
复制标题
DOI：
10.1091/mbc.e08-03-0297
复制
发表时间：
2008-10-01
影响因子：
3.3
通讯作者：
Simons, K. T.
中科院分区：
文献类型：
--
作者：
Cohen, R. L.;Espelin, C. W.;Simons, K. T.
Mif2p是哺乳动物着丝粒结合蛋白CENP - C在芽殖酵母中的同源蛋白。我们对酿酒酵母Mif2p的结构域进行了定位，并研究了其缺失后的表型结果。通过染色质免疫沉淀（ChIP）和电泳迁移率变动分析，我们进一步表明Mif2p结合在芽殖酵母着丝粒的CDEIII区域，可能与Ndc10p在空间上紧密相连。此外，ChIP实验表明Mif2p将大量的内、外着丝粒蛋白招募到酵母动粒上，但不包括Ndc80或Spc105复合物。我们已经确定了Mif2p的C末端二聚化结构域的晶体结构。它具有“cupin”折叠结构，在多肽链构象和二聚体几何形状上与一种细菌转录因子的二聚化结构域极其相似。Mif2p二聚体似乎是一种类似增强子体结构的一部分，该结构在芽殖酵母中对着丝粒组装起核心作用。
Mif2p is the budding-yeast orthologue of the mammalian centromere-binding protein CENP-C. We have mapped domains of Saccharomyces cerevisiae Mif2p and studied the phenotyptic consequences of their deletion. Using chromatin immunoprecipitation (ChIP) and electrophoretic mobility shift assays, we have further shown that Mif2p binds in the CDEIII region of the budding-yeast centromere, probably in close spatial association with Ndc10p. Moreover, ChIP experiments show that Mif2p recruits to yeast kinetochores a substantial subset of inner and outer kinetochore proteins, but not the Ndc80 or Spc105 complexes. We have determined the crystal structure of the C-terminal, dimerization domain of Mif2p. It has a "cupin" fold, extremely similar both in polypeptide chain conformation and in dimer geometry to the dimerization domain of a bacterial transcription factor. The Mif2p dimer seems to be part of an enhanceosome-like structure that nucleates kinetochore assembly in budding yeast.