Iwahara J, Kigawa T, Kitagawa K, Masumoto H, Okazaki T, Yokoyama S
Department of Biophysics and Biochemistry, Graduate School of Science, University of Tokyo, Bunkyo-ku, Tokyo 113.
EMBO J. 1998 Feb 2;17(3):827-37. doi: 10.1093/emboj/17.3.827.
CENP-B has been suggested to organize arrays of centromere satellite DNA into a higher order structure which then directs centromere formation and kinetochore assembly in mammalian chromosomes. The N-terminal portion of CENP-B is a 15 kDa DNA binding domain (DBD) consisting of two repeating units, RP1 and RP2. The DBD specifically binds to the CENP-B box sequence (17 bp) in centromere DNA. We determined the solution structure of human CENP-B DBD RP1 by multi-dimensional 1H, 13C and 15N NMR methods. The CENP-B DBD RP1 structure consists of four helices and has a helix-turn-helix structure. The overall folding is similar to those of some other eukaryotic DBDs, although significant sequence homology with these proteins was not found. The DBD of yeast RAP1, a telomere binding protein, is most similar to CENP-B DBD RP1. We studied the interaction between CENP-B DBD RP1 and the CENP-B box by the use of NMR chemical shift perturbation. The results suggest that CENP-B DBD RP1 interacts with one of the essential regions of the CENP-B box DNA, mainly at the N-terminal basic region, the N-terminal portion of helix 2 and helix 3.
有人提出，着丝粒蛋白B（CENP - B）可将着丝粒卫星DNA阵列组织成更高阶的结构，进而指导哺乳动物染色体中的着丝粒形成和动粒组装。CENP - B的N端部分是一个15 kDa的DNA结合结构域（DBD），由两个重复单元RP1和RP2组成。该DBD特异性结合着丝粒DNA中的CENP - B框序列（17 bp）。我们通过多维1H、13C和15N NMR方法确定了人CENP - B DBD RP1的溶液结构。CENP - B DBD RP1结构由四个螺旋组成，具有螺旋 - 转角 - 螺旋结构。尽管未发现与其他真核DBD有显著的序列同源性，但其整体折叠与其他一些真核DBD相似。酵母端粒结合蛋白RAP1的DBD与CENP - B DBD RP1最为相似。我们利用NMR化学位移扰动研究了CENP - B DBD RP1与CENP - B框之间的相互作用。结果表明，CENP - B DBD RP1与CENP - B框DNA的一个关键区域相互作用，主要位于N端碱性区域、螺旋2的N端部分和螺旋3。
