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Site‐Specific Investigation of the Steady‐State Kinetics and Dynamics of the Multistep Binding of Bile Acid Molecules to a Lipid Carrier Protein
Author(s) -
Cogliati Clelia,
Ragona Laura,
D'Onofrio Mariapina,
Günther Ulrich,
Whittaker Sara,
Ludwig Christian,
Tomaselli Simona,
Assfalg Michael,
Molinari Henriette
Publication year - 2010
Publication title -
chemistry – a european journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.687
H-Index - 242
eISSN - 1521-3765
pISSN - 0947-6539
DOI - 10.1002/chem.201000498
Subject(s) - ligand (biochemistry) , molecular dynamics , chemistry , macromolecule , biophysics , molecule , receptor–ligand kinetics , kinetics , plasma protein binding , binding site , mechanism (biology) , small molecule , stereochemistry , computational chemistry , biochemistry , receptor , biology , physics , philosophy , organic chemistry , epistemology , quantum mechanics
The investigation of multi‐site ligand–protein binding and multi‐step mechanisms is highly demanding. In this work, advanced NMR methodologies such as 2D 1 H– 15 N line‐shape analysis, which allows a reliable investigation of ligand binding occurring on micro‐ to millisecond timescales, have been extended to model a two‐step binding mechanism. The molecular recognition and complex uptake mechanism of two bile salt molecules by lipid carriers is an interesting example that shows that protein dynamics has the potential to modulate the macromolecule–ligand encounter. Kinetic analysis supports a conformational selection model as the initial recognition process in which the dynamics observed in the apo form is essential for ligand uptake, leading to conformations with improved access to the binding cavity. Subsequent multi‐step events could be modelled, for several residues, with a two‐step binding mechanism. The protein in the ligand‐bound state still exhibits a conformational rearrangement that occurs on a very slow timescale, as observed for other proteins of the family. A global mechanism suggesting how bile acids access the macromolecular cavity is thus proposed.