Mapping the encounter state of a transient protein complex by PRE NMR spectroscopy |
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Authors: | Alexander N Volkov Marcellus Ubbink Nico A J van Nuland |
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Institution: | (1) Department of Molecular and Cellular Interactions, VIB, Pleinlaan 2, 1050 Brussels, Belgium;(2) Jean Jeener NMR Centre, Structural Biology Brussels, Vrije Universiteit Brussel, Pleinlaan 2, 1050 Brussels, Belgium;(3) Leiden Institute of Chemistry, Gorlaeus Laboratories, Leiden University, P.O. Box 9502, 2300 RA Leiden, The Netherlands |
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Abstract: | Many biomolecular interactions proceed via a short-lived encounter state, consisting of multiple, lowly-populated species
invisible to most experimental techniques. Recent development of paramagnetic relaxation enhancement (PRE) nuclear magnetic
resonance (NMR) spectroscopy has allowed to directly visualize such transient intermediates in a number of protein-protein
and protein-DNA complexes. Here we present an analysis of the recently published PRE NMR data for a protein complex of yeast
cytochrome c (Cc) and cytochrome c peroxidase (CcP). First, we describe a simple, general method to map out the spatial and temporal distributions of binding
geometries constituting the Cc-CcP encounter state. We show that the spatiotemporal mapping provides a reliable estimate of
the experimental coverage and, at higher coverage levels, allows to delineate the conformational space sampled by the minor
species. To further refine the encounter state, we performed PRE-based ensemble simulations. The generated solutions reproduce
well the experimental data and lie within the allowed regions of the encounter maps, confirming the validity of the mapping
approach. The refined encounter ensembles are distributed predominantly in a region encompassing the dominant form of the
complex, providing experimental proof for the results of classical theoretical simulations. |
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