Myosin subfragment 1 hydrophobicity changes associated with different nucleotide-induced conformations.
Myosin subfragment 1 hydrophobicity changes associated with different nucleotide-induced conformations.
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肌球蛋白亚片段 1 疏水性变化与不同的核苷酸诱导构象相关。
DOI：
10.1021/bi951929c
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发表时间：
1996
期刊：
影响因子：
--
通讯作者：
Burke,M
中科院分区：
文献类型：
--
作者：
Gopal,D;Burke,M
肌球蛋白亚片段1的疏水性被认为是敏感的结合核苷酸在其活性位点的占用率和性质，如所示的未修饰和化学修饰的S1在苯基疏水色谱洗脱行为的变化。用N-乙基马来酰亚胺对SH 1（Cys-707）进行烷基化或用对苯二马来酰亚胺对SH 1和SH 2（Cys-697）进行共价桥连并捕获MgADP，均未改变S1的洗脱性能。虽然加入MgADP或MgATP的洗脱缓冲液对这些改性的S1物种的洗脱性能的影响最小，这些核苷酸的存在下，被发现与未改性的S1产生差异的影响。对于MgADP，其中S1处于S1*MgADP状态，洗脱时间略微减少，而对于MgATP，其中S1主要处于S1**MgADP·Pistate状态，洗脱时间显著降低，表明固定的苯基配体的可及性降低。稳定的S1三元复合物，形成MgADP和各种Pianalogues，显示洗脱时间类似于S1在含有MgATP的缓冲液。因此，两个主要类别的核苷酸诱导的S1构象可以根据它们与固定化苯基的相互作用来定义。S1疏水性的这些核苷酸诱导的变化与报道的与结合核苷酸的不同状态相关的S1回转半径的变化良好相关[Wakabayashi，K.，Tokunga，M.，科诺岛Sugimoto，Y.; Hamanaka，T.，Takezawa，Y.，Wakabayashi，T.，& Amemiya，Y.（1992）Science 258，443 - 447]，表明观察到的疏水性相互作用可能是测量固定的苯基配体进入疏水缝隙的可接近性，并且当S1处于S1**MgADP·Pistate中时，该缝隙闭合或收紧。
Myosin subfragment 1 hydrophobicity was found to be sensitive to the occupancy and nature of bound nucleotide at its active site, as shown by changes in elution behavior of unmodified and chemically modified S1 during phenyl hydrophobic chromatography. The elution properties of S1 were unaltered by alkylation of SH1 (Cys-707) withN-ethylmaleimide or by covalent bridging between SH1 and SH2 (Cys-697) withp-phenylenedimaleimide with trapping of MgADP. Although addition of MgADP or MgATP to the elution buffers had minimal effect on the elution properties of these modified S1 species, the presence of these nucleotides was found to produce differential effects with unmodified S1. With MgADP, where S1 is in the S1*MgADP state, the elution times were decreased slightly, whereas with MgATP, where S1 is primarily in the S1**MgADP·Pistate, the elution times were significantly lowered, indicating reduced accessibility for the immobilized phenyl ligand. Stable S1 ternary complexes, formed with MgADP and various Pianalogues, showed elution times similar to that for S1 in the buffers containing MgATP. Thus, two main classes of nucleotide-induced S1 conformations can be defined according to their interaction with immobilized phenyl. These nucleotide-induced changes in S1 hydrophobicity correlate well with reported changes in radius of gyration of S1 associated with different states of the bound nucleotide [Wakabayashi, K., Tokunga, M., Kohno, I., Sugimoto, Y.; Hamanaka, T., Takezawa, Y., Wakabayashi, T., & Amemiya, Y. (1992)Science258, 443−447], suggesting that the observed hydrophobicity interaction may be measuring accessibility of the immobilized phenyl ligand into a hydrophobic crevice, and that this crevice is closed or tightened when S1 is in the S1**MgADP·Pistate.