A human centromere protein, CENP-B, has a DNA binding domain containing four potential a helices at the NH2 terminus, which is separable from dimerizing activity

Kinya Yoda, Katsumi Kitagawa, Hiroshi Masumoto, Yoshinao Muro, Tuneko Okazaki

Research output: Contribution to journalArticle

102 Scopus citations

Abstract

The alphoid DNA-CENP-B (centromere protein B) complex is the first sequence-specific DNA/protein complex detected in the centromeric region of human chromosomes. In the reaction, CENP-B recognizes a 17-bp sequence (CENP-B box) and assembles two alphoid DNA molecules into a complex, which is designated complex A (Muro, Y., H. Masumoto, K. Yoda, N. Nozaki, M. Ohashi, and T. Okazaki. 1992. J. Cell Biol. 116:585-596). Since CENP-B gene is conserved in mammalian species and CENP-B boxes are found also in mouse centromere satellite DNA (minor satellite), this sequence-specific DNA-protein interaction may be important for some kind of common centromere function. In this study we have characterized the structure of CENP-B and CENP-B-alphoid DNA complex. We have shown by chemical cross-linking that CENP-B formed a dimer, and have estimated by molecular weight determination the composition of complex A to be a CENP-B dimer and two molecules of alphoid DNA. The DNA binding domain has been delimited within the NH2-terminal 125-amino acid region containing four potential α-helices using truncated CENP-B made in Escherichia coli cells. We have shown that CENP-B had sites highly sensitive to proteases and that the DNA binding domain was separable from the dimerizing activity by the proteolytic cleavage at 20 kD from the COOH terminus of the molecule. Thus, CENP-B may organize a higher order structure in the centromere by juxtaposing two CENP-B boxes in the alphoid DNA repeat through both the DNA-protein and protein-protein interactions.

Original languageEnglish (US)
Pages (from-to)1413-1427
Number of pages15
JournalJournal of Cell Biology
Volume119
Issue number6
DOIs
StatePublished - Dec 1992
Externally publishedYes

ASJC Scopus subject areas

  • Cell Biology

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