Single-molecule observation of the induction of k-turn RNA structure on binding L7Ae protein

Jia Wang, Tomáš Fessl, Kersten T. Schroeder, Jonathan Ouellet, Yijin Liu, Alasdair D. J. Freeman, David M. J. Lilley (Lead / Corresponding author)

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    25 Citations (Scopus)
    120 Downloads (Pure)

    Abstract

    The k-turn is a commonly occurring structural motif that introduces a tight kink into duplex RNA. In free solution, it can exist in an extended form, or by folding into the kinked structure. Binding of proteins including the L7Ae family can induce the formation of the kinked geometry, raising the question of whether this occurs by passive selection of the kinked structure, or a more active process in which the protein manipulates the RNA structure. We have devised a single-molecule experiment whereby immobilized L7Ae protein binds Cy3-Cy5-labeled RNA from free solution. We find that all bound RNA is in the kinked geometry, with no evidence for transitions to an extended form at the millisecond timescale of the camera. Furthermore, real-time binding experiments provide no evidence for a more extended intermediate even at the earliest times, at a time resolution of 16 ms. The data support a passive conformational selection model by which the protein selects a fraction of RNA that is already in the kinked conformation, thereby drawing the equilibrium into this form.

    Original languageEnglish
    Pages (from-to)2541-2548
    Number of pages8
    JournalBiophysical Journal
    Volume103
    Issue number12
    Early online date18 Dec 2012
    DOIs
    Publication statusPublished - 19 Dec 2012

    ASJC Scopus subject areas

    • Biophysics

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