Peptide-Protein Binding Investigated by Far-IR Spectroscopy and Molecular Dynamics Simulations

Abstract

Molecular dynamics (MD) simulations and far-infrared (far-IR) spectroscopy were combined to study peptide binding by the second PDZ domain (PDZ1) of MAGI1, which has been identified as an important target for the Human Papilloma Virus. PDZ1 recognizes and binds to the C-terminal end of the E6 protein from high-risk Human Papilloma Virus. The far-IR spectra of two forms of the protein, an unbound APO form and a HOLO form (where the PDZ1 is bound to an 11-residue peptide derived from the C terminus of HPV16 E6), were obtained. MD simulations were used to determine the most representative structure of each form and these were used to compute their respective IR spectra by normal mode analysis. Far-UV circular dichroism spectroscopy was used to confirm the secondary structure content and the stability through temperature-dependent studies. Both the experimental and calculated far-IR spectra showed a red shift of the low-frequency peaks upon peptide binding. The calculations show that this is coincident with an increased number of hydrogen bonds formed as the peptide augments the protein β-sheet. We further identified the contribution of surface-bound water molecules to bands in the far-IR and, through the calculations, identified potential pathways for allosteric communication. Together, these results demonstrate the utility of combining far-IR experiments and MD studies to study peptide binding by proteins.

Persistent Protein Motions in a Rugged Energy Landscape Revealed by Normal Mode Ensemble Analysis

“…For these studies, we use the benchmark protein hen egg white lysozyme (HEWL), for which there are extensive measurements of picosecond dynamics using a wide variety of techniques and whose results are representative of other proteins. Neutron scattering, Raman, Optical Kerr, and isotropic terahertz spectroscopy have found broad vibrational density of states and isotropic absorbance, with only slight changes in this absorbance with functional state. , However, recent terahertz anisotropic absorption studies on HEWL find narrow band resonances that are sensitive to inhibitor binding. , By rigorously averaging spectra calculated from structures sampled from microsecond-scale all-atom trajectories, we find that indeed the averaged isotropic spectra do not have narrow resonances and resemble the smooth absorbance seen experimentally. On the other hand, resonant bands are present in the average anisotropic absorbance spectra.…”

“…Neutron scattering, Raman, Optical Kerr, and isotropic terahertz spectroscopy have found broad vibrational density of states and isotropic absorbance, with only slight changes in this absorbance with functional state. 14,15 However, recent terahertz anisotropic absorption studies on HEWL find narrow band resonances that are sensitive to inhibitor binding. 16,17 By rigorously averaging spectra calculated from structures sampled from microsecond-scale all-atom trajectories, we find that indeed the averaged isotropic spectra do not have narrow resonances and resemble the smooth absorbance seen experimentally.…”

Protein and RNA dynamical fingerprinting

“…While careful and precise isotropic absorbance measurements have detected changes in the broad absorption peaks with substrate/inhibitor binding for a variety of proteins 39,40 , the polarization dependent THz absorption methods used here remove the isotropic water absorption and increase detection selectivity, isolating resonant bands based on the direction of the transition dipoles. It is notable that the near field measurements reflect the net absorbance for a large population of biomolecules which are distributed over a rugged energy landscape.…”

Investigating Membrane‐Mediated Antimicrobial Peptide Interactions with Synchrotron Radiation Far‐Infrared Spectroscopy

“…[17,18] Supplemented with 2D correlative analysis, FTIR becomes a powerful tool to put into evidence the potential interdependencies and interactions between peptides and their (native) molecular environments. Complementary to the mid-IR region (MIR), the use of the low frequency region, far-FTIR (FIR) [19] or terahertz (THz)-time domain [20] is intrinsically attractive as it discriminates vibrational modes involving interand intramolecular hydrogen bonding [21] which, due to their collective nature, are highly sensitive to the conformational state of the molecule. [22] Such a capability can then provide a unique fingerprint for a specific molecular arrangement.…”