Identification of Strategic Residues at the Interface of Antigen–Antibody Interactions by In Silico Mutagenesis

Abstract

Structural information pertaining to antigen-antibody interactions is fundamental in immunology, and benefits structure-based vaccine design. Modeling of antigen-antibody immune complexes from co-crystal structures or molecular docking simulations provides an extensive profile of the epitope at the interface; however, the key amino acids involved in the interaction must be further clarified, often through the use of experimental mutagenesis and subsequent binding assays. Here, we describe an in silico mutagenesis method to identify key sites at antigen-antibody interfaces, using significant increase in pH-dependency energy among saturated point mutations. Through a comprehensive analysis of the crystal structures of three antigen-antibody immune complexes, we show that a cutoff value of 1 kcal/mol of increased interaction energy provides good congruency with the experimental non-binding mutations conducted in vitro. This in silico mutagenesis strategy, in association with energy calculations, may provide an efficient tool for antibody-antigen interface analyses, epitope optimization, and/or conformation prediction in structure-based vaccine design.

Binding Modes of Small‐Molecule Inhibitors to the EED Pocket of PRC2

“…[39][40][41][42] The structure of EED was taken from the EED/EED226 complex structure (PDB ID: 5GSA). [39][40][41][42] The structure of EED was taken from the EED/EED226 complex structure (PDB ID: 5GSA).…”

“…[33,34,42] Water molecules in the crystal structure were preserved. [33,34,42] Water molecules in the crystal structure were preserved.…”

“…The geometry of ligand MAK683 was optimized using the DFT (B3LYP) method and 6-31 g + basis set implemented in Gaussian 09 and then employed in the molecular docking. [39][40][41][42] The structure of EED was taken from the EED/EED226 complex structure (PDB ID: 5GSA). AutoDock4 was used for proteinligand docking of EED with ligand MAK683.…”

“…Three EED/ligand complexes, EED/EED226 (PDB ID: 5GSA), EED/ A-395 (PDB ID: 5K0M), and the docked EED/MAK683 structure are subjected to MD simulation and computational alanine scanning. [33,34,42] Water molecules in the crystal structure were preserved. For each system, missing atoms were added using the tleap module of AMBER14 suite and a truncated octahedral TIP3P water box with a buffer distance of 12.0 Å was used to solvate the complex.…”

Molecular Dynamics Simulations in Designing DARPins as Phosphorylation-Specific Protein Binders of ERK2

“…These methods are fast, user-friendly, and reliable and promise to be invaluable in the development of proteins with a wide range of impactful applications. The applications of these tools extend from understanding the origin of diseases caused by misregulation of protein maintenance [ 51 , 52 ] to discriminating disease-associated mutations from non-disease mutations, studying drug-resistant mutations [ 53 , 54 , 55 ], and providing important structural and functional insights into designing new proteins [ 56 , 57 , 58 , 59 ]. To design DARPins as ERK2 inhibitors, a multi-point mutation approach, MAESTRO, was applied to the wild-type DARPin protein to identify the stabilizing mutation points, followed by validation of the binding energy of mutants employing MD simulations using MM-PBSA/GBSA protocols.…”