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Characterization of monobody scaffold interactions with ligand via force spectroscopy and steered molecular dynamics.

ABSTRACT: Monobodies are antibody alternatives derived from fibronectin that are thermodynamically stable, small in size, and can be produced in bacterial systems. Monobodies have been engineered to bind a wide variety of target proteins with high affinity and specificity. Using alanine-scanning mutagenesis simulations, we identified two scaffold residues that are critical to the binding interaction between the monobody YS1 and its ligand, maltose-binding protein (MBP). Steered molecular dynamics (SMD) simulations predicted that the E47A and R33A mutations in the YS1 scaffold substantially destabilize the YS1-MBP interface by reducing the bond rupture force and the lifetime of single hydrogen bonds. SMD simulations further indicated that the R33A mutation weakens the hydrogen binding between all scaffold residues and MBP and not just between R33 and MBP. We validated the simulation data and characterized the effects of mutations on YS1-MBP binding by using single-molecule force spectroscopy and surface plasmon resonance. We propose that interfacial stability resulting from R33 of YS1 stacking with R344 of MBP synergistically stabilizes both its own bond and the interacting scaffold residues of YS1. Our integrated approach improves our understanding of the monobody scaffold interactions with a target, thus providing guidance for the improved engineering of monobodies.

SUBMITTER: Cheung LS

PROVIDER: S-EPMC4316159 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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Characterization of monobody scaffold interactions with ligand via force spectroscopy and steered molecular dynamics.

Cheung Luthur Siu-Lun LS Shea Daniel J DJ Nicholes Nathan N Date Amol A Ostermeier Marc M Konstantopoulos Konstantinos K

Scientific reports 20150204

Monobodies are antibody alternatives derived from fibronectin that are thermodynamically stable, small in size, and can be produced in bacterial systems. Monobodies have been engineered to bind a wide variety of target proteins with high affinity and specificity. Using alanine-scanning mutagenesis simulations, we identified two scaffold residues that are critical to the binding interaction between the monobody YS1 and its ligand, maltose-binding protein (MBP). Steered molecular dynamics (SMD) si ...[more]

PMID: 25650239

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Project description:Computing the free energy of binding a ligand to a protein is a difficult task of essential importance for which purpose various theoretical/computational approaches have been pursued. In this paper, we develop a hybrid steered molecular dynamics (hSMD) method capable of resolving one ligand–protein complex within a few wall-clock days with high enough accuracy to compare with the experimental data. This hSMD approach is based on the relationship between the binding affinity and the potential of mean force (PMF) in the established literature. It involves simultaneously steering n (n = 1, 2, 3, ...) centers of mass of n selected segments of the ligand using n springs of infinite stiffness. Steering the ligand from a single initial state chosen from the bound state ensemble to the corresponding dissociated state, disallowing any fluctuations of the pulling centers along the way, one can determine a 3n-dimensional PMF curve connecting the two states by sampling a small number of forward and reverse pulling paths. This PMF constitutes a large but not the sole contribution to the binding free energy. Two other contributors are (1) the partial partition function containing the equilibrium fluctuations of the ligand at the binding site and the deviation of the initial state from the PMF minimum and (2) the partial partition function containing rotation and fluctuations of the ligand around one of the pulling centers that is fixed at a position far from the protein. We implement this hSMD approach for two ligand–protein complexes whose structures were determined and whose binding affinities were measured experimentally: caprylic acid binding to bovine ?-lactoglobulin and glutathione binding to Schistosoma japonicum glutathione S-transferase tyrosine 7 to phenylalanine mutant. Our computed binding affinities agree with the experimental data within a factor of 1.5. The total time of computation for these two all-atom model systems (consisting of 96K and 114K atoms, respectively) was less than one wall-clock week using 512 cores (32 Xeon E5-2680 processors).

Dataset Information

Characterization of monobody scaffold interactions with ligand via force spectroscopy and steered molecular dynamics.

Publications

Characterization of monobody scaffold interactions with ligand via force spectroscopy and steered molecular dynamics.

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