Combining Free Energy Simulations and NMR Chemical-Shift Perturbation To Identify Transient Cation−π Contacts in Proteins

Abstract

Flexible protein regions containing cationic and aromatic side-chains exposed to solvent may form transient cation-π interactions with structural and functional roles. To evaluate their stability and identify important intramolecular cation-π contacts, a combination of free energy profiles estimated from umbrella sampling with molecular dynamics simulations and chemical shift perturbations (CSP) obtained from NMR experiments is applied here to the complete catalytic domain of human phosphatase Cdc25B. This protein is a good model system for transient cation-π interactions as it contains only one Trp residue (W550) in the disordered Cterminal segment and a total of 17 Arg residues, many exposed to solvent. Eight putative Arg-Trp pairs were simulated here. Only R482 and R544 show bound profiles corresponding to important transient cation-π interactions, while the others have dissociative or almost flat profiles. These results are corroborated by CSP analysis of three Cdc25B point mutants (W550A, R482A and R544A) disrupting cation-π contacts. The proposed validation of statistically representative molecular simulations by NMR spectroscopy could be applied to identify transient contacts of proteins in general but carefully, as NMR chemical shifts are sensitive to changes in both molecular contacts and conformational distributions.

Mechanism of rotenone binding to respiratory complex I depends on ligand flexibility

“…A rotenone molecule bound to ROT2 was only built in open state cryoEM models (PDBs 6ZKL and 6ZKM) 8 . The NDUFS7 Arg87 loop in these open-bound models is also reshaped to a closed conformation 8 , 15 , disrupting a cation- stacking 58 found between Arg87 and Phe86 in open-unbound models and placing Arg87 towards the Q-channel entrance (Fig. 3 B).…”

Mechanisms for Flavin-Mediated Oxidation: Hydride or Hydrogen-Atom Transfer?

“…1,2 The redox center in all natural flavins is formed by the heteronuclear tricyclic isoallox-azine ring (Figure 1), primarily attached to the protein by noncovalent hydrogen bonds, stacking and cation-π contacts. [3][4][5] These interactions also modulate the flavin redox potential from −400 to 60 mV, allowing oxidation of a range of aliphatic and aromatic substrates. [6][7][8] Flavin redox reactions are an example of proton-coupled electron transfers or PCET, a broad family of reactions and energy conversion processes in chemistry.…”

“…Enzymes equipped with flavin cofactors comprise the most abundant class of natural catalysts for combined proton and electron transfer. , The redox center in all natural flavins is formed by the heteronuclear tricyclic isoalloxazine ring (Figure ), primarily attached to the protein by noncovalent hydrogen bonds, stacking, and cation−π contacts. − These interactions also modulate the flavin redox potential from −400 to 60 mV, allowing oxidation of a range of aliphatic and aromatic substrates. − …”

Accuracy of Alternate Nonpolarizable Force Fields for the Determination of Protein–Ligand Binding Affinities Dominated by Cation−π Interactions

“…Protein–ligand association plays a pivotal role in a wide range of biological processes. , Absolute binding free energy calculations based on a rigorous framework have proven particularly useful to predict up to chemical accuracy the propensity of a ligand to associate to a protein in silico . − Although the results obtained from such free-energy calculations are usually close to the experimental measurements, − nontrivial errors may sometimes deteriorate the agreement, which is believed to stem, at least in part, from unusual noncovalent interactions, which are not always well described by the adopted macromolecular force field, such as cation−π, salt–bridge, and π–π interactions. ,, In general, users rely on the default parameters of popular academic force fields, such as CHARMM, − AMBER, , and OPLS, , without delving too much on the quality of the description of these unusual noncovalent interactions. Of particular concern are cation−π interactions, , a considerable number of which can been found in protein-based objects through structural analysis of the Protein Data Bank­(PDB). − The dominating contributions in cation−π interactions arise from two different effects. The electric field generated by the cation polarizes the π–electron cloud of the aromatic ring, and, in turn, the induced dipole of the aromatic ring interacts with the polarizing charge. , From a physics standpoint, both contributions vary in r –2 , resulting in an attractive part of the potential that describes induction effects varying in r –4 . , Not too surprisingly, cation−π interactions have been copiously reported as being poorly represented by the default parameters of popular force fields, especially when quantifying interaction energies. − …”