• 我要登录|
  • 免费注册
    |
  • 我的丁香通
    • 企业机构:
    • 成为企业机构
    • 个人用户:
    • 个人中心
  • 移动端
    移动端
丁香通 logo丁香实验_LOGO
搜实验

    大家都在搜

      大家都在搜

        0 人通过求购买到了急需的产品
        免费发布求购
        发布求购
        点赞
        收藏
        wx-share
        分享

        Quantification of Poly(ADP-Ribose) In Vitro: Determination of the ADP-Ribose Chain Length and Branching Pattern

        互联网

        466
        The structural integrity of eukaryotic genomes, to a great extent, depends on highly regulated and �coordinated enzymatic chromosomal poly(ADP-ribosyl)ation cycles that target chromatin proteins for specific covalent epigenetic poly(ADP-ribose) modification. As a result, the accurate determination of poly(ADP-ribosyl)ation amino acid specificity, as well as, a detailed characterization of the structural �complexity of the protein-bound ADP-ribose polymers generated, e.g., linear versus branched ADP-ribose chains, need to be carefully sorted out. In this chapter, we describe well-established and reproducible laboratory methods and protocols typically used to determine: (1) the ADP-ribose chain length(s) and (2) the molecular stoichiometry of the protein–poly(ADP-ribosyl)ation reaction, e.g., number of ADP-ribose chains/polypeptide unit. While the methodology described here is exclusively for in vitro purified systems that can be used with high reliability, the reader is advised that application of these protocols to whole cell extracts and tissue systems must take into consideration the rapid turnover rate of protein-bound ADP-ribose polymers in vivo. Indeed, these extremely low-abundance chromatin-bound polymeric molecules are notoriously characterized for displaying a short half-life, typically from a few seconds to a few minutes. We also discuss potential methodological pitfalls, such as: (1) the chemical stability of protein–(ADP-ribose)n adducts and (2) the requirement for polymeric radiolabeling.
        ad image
        提问
        扫一扫
        丁香实验小程序二维码
        实验小助手
        丁香实验公众号二维码
        扫码领资料
        反馈
        TOP
        打开小程序