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Single-molecule analysis of Mss116 -mediated group II intron folding

机译：Mss116介导的II组内含子折叠的单分子分析

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摘要

DEAD-box helicases are conserved enzymes involved in nearly all aspects of RNA metabolism, but their mechanisms of action remain unclear. Here, we investigated the mechanism of the DEAD-box protein Mss116 on its natural substrate, the group II intron ai5γ. Group II introns are structurally complex catalytic RNAs considered evolutionarily related to the eukaryotic spliceosome, and an interesting paradigm for large RNA folding. We used single-molecule fluorescence to monitor the effect of Mssll6 on folding dynamics of a minimal active construct, ai5y-D135. The data show that Mssll6 stimulates dynamic sampling between states along the folding pathway, an effect previously observed only with high Mg~(2+) concentrations. Furthermore, the data indicate that Mssll6 promotes folding through discrete ATP-independent and ATP-dependent steps. We propose that Mssll6 stimulates group II intron folding through a multi-step process that involves electrostatic stabilization of early intermediates and ATP hydrolysis during the final stages of native state assembly.
机译：DEAD-box解旋酶是保守的酶，几乎涉及RNA代谢的所有方面，但其作用机理仍不清楚。在这里，我们研究了DEAD-box蛋白Mss116在其天然底物II组内含子ai5γ上的作用机理。 II组内含子是结构复杂的催化RNA，被认为与真核剪接体进化相关，并且是大RNA折叠的有趣范例。我们使用单分子荧光来监测Mssll6对最小活性构建体ai5y-D135折叠动力学的影响。数据显示，Mssll6刺激沿折叠路径的状态之间的动态采样，以前只有在高Mg〜（2+）浓度下才能观察到这种效果。此外，数据表明Mssll6通过不依赖ATP和不依赖ATP的步骤促进折叠。我们建议，Mssll6通过多步过程刺激II组内含子折叠，该过程涉及早期中间体的静电稳定和天然状态组装最后阶段的ATP水解。

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  • 来源
    《Nature》 |2010年第7318期|P.935-939|共5页
  • 作者

    Krishanthi S. Karunatilaka; Amanda Solem; Anna Marie Pyle; David Rueda;

  • 作者单位

    Department of Chemistry, Wayne State University, 5101 Cass Avenue, Detroit, Michigan 48202, USA;

    rnDepartment of Chemistry, Wayne State University, 5101 Cass Avenue, Detroit, Michigan 48202, USA;

    rnDepartment of Molecular, Cellular and Developmental Biology and Department of Chemistry, Yale University, New Haven, Connecticut 06520, USA Howard Hughes Medical Institute, Yale University, New Haven, Connecticut 06520, USA;

    rnDepartment of Chemistry, Wayne State University, 5101 Cass Avenue, Detroit, Michigan 48202, USA;

  • 收录信息 美国《科学引文索引》(SCI);美国《工程索引》(EI);美国《生物学医学文摘》(MEDLINE);美国《化学文摘》(CA);
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