Addressing Challenges of Macrocyclic Conformational Sampling in Polar and Apolar Solvents: Lessons for Chameleonicity.

We evaluated a workflow to reliably sample the conformational space of a set of 47 peptidic macrocycles. Starting from SMILES strings, we use accelerated molecular dynamics simulations to overcome high energy barriers, in particular, the cis-trans isomerization of peptide bonds. We find that our approach performs very well in polar solvents like water and dimethyl sulfoxide.

Structure and Dynamics Guiding Design of Antibody Therapeutics and Vaccines.

Antibodies and other new antibody-like formats have emerged as one of the most rapidly growing classes of biotherapeutic proteins. Understanding the structural features that drive antibody function and, consequently, their molecular recognition is critical for engineering antibodies. Here, we present the structural architecture of conventional IgG antibodies alongside other formats.

Influence of the Fluorophore Mobility on Distance Measurements by Gas-Phase FRET.

Gas-phase Förster resonance energy transfer (FRET) combines mass spectrometry and fluorescence spectroscopy for the conformational analysis of mass-selected biomolecular ions. In FRET, fluorophore pairs are typically covalently attached to a biomolecule using short linkers, which affect the mobility of the dye and the relative orientation of the transition dipole moments of the donor and acceptor. Intramolecular interactions may further influence the range of motion.

Determining the gas-phase structures of α-helical peptides from shape, microsolvation, and intramolecular distance data.

Mass spectrometry is a powerful technique for the structural and functional characterization of biomolecules. However, it remains challenging to accurately gauge the gas-phase structure of biomolecular ions and assess to what extent native-like structures are maintained. Here we propose a synergistic approach which utilizes Förster resonance energy transfer and two types of ion mobility spectrometry (i.

Lessons for Oral Bioavailability: How Conformationally Flexible Cyclic Peptides Enter and Cross Lipid Membranes.

Cyclic peptides extend the druggable target space due to their size, flexibility, and hydrogen-bonding capacity. However, these properties impact also their passive membrane permeability. As the "journey" through membranes cannot be monitored experimentally, little is known about the underlying process, which hinders rational design.

Covalent polyphenol modification of a reactive cysteine in the major apple allergen Mal d 1.

Naturally occurring polyphenols can modify the molecular properties of food allergens. For the major apple allergen Mal d 1 it has been postulated that chemical reactions with polyphenols cause permanent changes in the tertiary structure, causing a loss of conformational IgE epitopes and reducing allergenicity. In our study, we investigated the effect that reactions with oxidized polyphenols have on the structure of Mal d 1 by mass spectrometry and NMR spectroscopy.

pH-dependent structural diversity of profilin allergens determines thermal stability.

The family of profilin allergens is a common class of proteins found in plants, viruses and various eukaryotes including mammals. Profilins are characterized by an evolutionary conserved structural fold, which is responsible for their cross-reactive nature of Immunoglobulin E (IgE) antibodies. Despite their high overall structural similarity, they exhibit substantial differences in their biophysical properties, such as thermal and pH stability.

Ascorbylation of a Reactive Cysteine in the Major Apple Allergen Mal d 1.

The protein Mal d 1 is responsible for most allergic reactions to apples () in the northern hemisphere. Mal d 1 contains a cysteine residue on its surface, with its reactive side chain thiol exposed to the surrounding food matrix. We show that, in vitro, this cysteine residue is prone to spontaneous chemical modification by ascorbic acid (vitamin C).

Grid inhomogeneous solvation theory for cross-solvation in rigid solvents.

Grid Inhomogeneous Solvation Theory (GIST) has proven useful to calculate localized thermodynamic properties of water around a solute. Numerous studies have leveraged this information to enhance structure-based binding predictions. We have recently extended GIST toward chloroform as a solvent to allow the prediction of passive membrane permeability.

Enhanced sampling without borders: on global biasing functions and how to reweight them.

Molecular dynamics (MD) simulations are a powerful tool to follow the time evolution of biomolecular motions in atomistic resolution. However, the high computational demand of these simulations limits the timescales of motions that can be observed. To resolve this issue, so called enhanced sampling techniques are developed, which extend conventional MD algorithms to speed up the simulation process.

Energy penalties enhance flexible receptor docking in a model cavity.

Protein flexibility remains a major challenge in library docking because of difficulties in sampling conformational ensembles with accurate probabilities. Here, we use the model cavity site of T4 lysozyme L99A to test flexible receptor docking with energy penalties from molecular dynamics (MD) simulations. Crystallography with larger and smaller ligands indicates that this cavity can adopt three major conformations: open, intermediate, and closed.

Structure and Zeatin Binding of the Peach Allergen .

Peach () is among the fruits most frequently reported to cause food allergies. Allergic reactions commonly result from previous sensitization to the birch pollen allergen Bet v 1, followed by immunological cross-reactivity of IgE antibodies to structurally related proteins in peach. In this study, we present the three-dimensional NMR solution structure of the cross-reactive peach allergen (isoform Pru p 1.

X-Entropy: A Parallelized Kernel Density Estimator with Automated Bandwidth Selection to Calculate Entropy.

is a Python package used to calculate the entropy of a given distribution, in this case, based on the distribution of dihedral angles. The dihedral entropy facilitates an alignment-independent measure of local protein flexibility. The key feature of our approach is a Gaussian kernel density estimation (KDE) using a plug-in bandwidth selection, which is fully implemented in a C++ backend and parallelized with OpenMP.

Inverse relation between structural flexibility and IgE reactivity of Cor a 1 hazelnut allergens.

A major proportion of allergic reactions to hazelnuts (Corylus avellana) are caused by immunologic cross-reactivity of IgE antibodies to pathogenesis-related class 10 (PR-10) proteins. Intriguingly, the four known isoforms of the hazelnut PR-10 allergen Cor a 1, denoted as Cor a 1.0401-Cor a 1.

pH-Induced Local Unfolding of the Phl p 6 Pollen Allergen From cpH-MD.

Susceptibility to endosomal degradation is a decisive contribution to a protein's immunogenicity. It is assumed that the processing kinetics of structured proteins are inherently linked to their probability of local unfolding. In this study, we quantify the impact of endosomal acidification on the conformational stability of the major timothy grass pollen allergen Phl p 6.

Conformational Ensembles of Antibodies Determine Their Hydrophobicity.

A major challenge in the development of antibody biotherapeutics is their tendency to aggregate. One root cause for aggregation is exposure of hydrophobic surface regions to the solvent. Many current techniques predict the relative aggregation propensity of antibodies via precalculated scales for the hydrophobicity or aggregation propensity of single amino acids.

Polarizable and non-polarizable force fields: Protein folding, unfolding, and misfolding.

Molecular dynamics simulations are an invaluable tool to characterize the dynamic motions of proteins in atomistic detail. However, the accuracy of models derived from simulations inevitably relies on the quality of the underlying force field. Here, we present an evaluation of current non-polarizable and polarizable force fields (AMBER ff14SB, CHARMM 36m, GROMOS 54A7, and Drude 2013) based on the long-standing biophysical challenge of protein folding.

Thermosensitive Hydration of Four Acrylamide-Based Polymers in Coil and Globule Conformations.

To characterize the thermosensitive coil-globule transition in atomistic detail, the conformational dynamics of linear polymer chains of acrylamide-based polymers have been investigated at multiple temperatures. Therefore, molecular dynamic simulations of 30mers of polyacrylamide (AAm), poly--methylacrylamide (NMAAm), poly--ethylacrylamide (NEAAm), and poly--isopropylacrylamide (NIPAAm) have been performed at temperatures ranging from 250 to 360 K for 2 μs. While two of the polymers are known to exhibit thermosensitivity (NEAAm, NIPAAm), no thermosensitivity is observed for AAm and NMAAm in aqueous solution.

Design of Phl p 6 Variants With Altered Fold-Stability Significantly Impacts Antigen Processing, Immunogenicity and Immune Polarization.

Understanding, which factors determine the immunogenicity and immune polarizing properties of proteins, is an important prerequisite for designing better vaccines and immunotherapeutics. While extrinsic immune modulatory factors such as pathogen associated molecular patterns are well-understood, far less is known about the contribution of protein inherent features. Protein fold-stability represents such an intrinsic feature contributing to immunogenicity and immune polarization by influencing the amount of peptide-MHC II complexes (pMHCII).

Conformational selection of allergen-antibody complexes-surface plasticity of paratopes and epitopes.

Antibodies have the ability to bind various types of antigens and to recognize different antibody-binding sites (epitopes) of the same antigen with different binding affinities. Due to the conserved structural framework of antibodies, their specificity to antigens is mainly determined by their antigen-binding site (paratope). Therefore, characterization of epitopes in combination with describing the involved conformational changes of the paratope upon binding is crucial in understanding and predicting antibody-antigen binding.

Profiling selectivity of chagasin mutants towards cysteine proteases cruzain or cathepsin L through molecular dynamics simulations.

Chagasin, an endogenous cysteine protease inhibitor from , can control the activity of the parasitic cruzain and its homologous human cathepsin L. While chagasin inhibits both enzymes with similar potency, mutations have different effects on binding to these enzymes. Mutants T31A and T31A/T32A bind well to cathepsin L, but their affinity for cruzain drops ∼40 to 140-fold.

Protein-Protein Binding as a Two-Step Mechanism: Preselection of Encounter Poses during the Binding of BPTI and Trypsin.

Biomolecular recognition between proteins follows complex mechanisms, the understanding of which can substantially advance drug discovery efforts. Here, we track each step of the binding process in atomistic detail with molecular dynamics simulations using trypsin and its inhibitor bovine pancreatic trypsin inhibitor (BPTI) as a model system. We use umbrella sampling to cover a range of unbinding pathways.

Solvation Thermodynamics in Different Solvents: Water-Chloroform Partition Coefficients from Grid Inhomogeneous Solvation Theory.

Reliable information on partition coefficients plays a key role in drug development, as solubility decisively affects bioavailability. In a physicochemical context, the partition coefficient of a solute between two different solvents can be described as a function of solvation free energies. Hence, substantial scientific efforts have been made toward accurate predictions of solvation free energies in various solvents.

Macrocycle Cell Permeability Measured by Solvation Free Energies in Polar and Apolar Environments.

The relation of surface polarity and conformational preferences is decisive for cell permeability and thus bioavailability of macrocyclic drugs. Here, we employ grid inhomogeneous solvation theory (GIST) to calculate solvation free energies for a series of six macrocycles in water and chloroform as a measure of passive membrane permeability. We perform accelerated molecular dynamics simulations to capture a diverse structural ensemble in water and chloroform, allowing for a direct profiling of solvent-dependent conformational preferences.

Catalytic Site p Values of Aspartic, Cysteine, and Serine Proteases: Constant pH MD Simulations.

Enzymatic function and activity of proteases is closely controlled by the pH value. The protonation states of titratable residues in the active site react to changes in the pH value, according to their p, and thereby determine the functionality of the enzyme. Knowledge of the titration behavior of these residues is crucial for the development of drugs targeting the active site residues.