Synthesizing Amino Acids Modified with Reactive Carbonyls in Silico to Assess Structural Effects Using Molecular Dynamics Simulations

Summary

Here, we describe a protocol for the optimization and parameterization of amino acid residues modified with reactive carbonyl species, adaptable to protein systems. The protocol steps include structure design and optimization, charge assignments, parameter construction, and preparation of protein systems.

Abstract

Protein carbonylation by reactive aldehydes derived from lipid peroxidation leads to cross-linking, oligomerization, and aggregation of proteins, causing intracellular damage, impaired cell functions, and, ultimately, cell death. It has been described in aging and several age-related chronic conditions. However, the basis of structural changes related to the loss of function in protein targets is still not well understood. Hence, a route to the in silico construction of new parameters for amino acids carbonylated with reactive carbonyl species derived from fatty acid oxidation is described. The Michael adducts for Cys, His, and Lys with 4-hydroxy-2-nonenal (HNE), 4-hydroxy-2-hexenal (HHE), and a furan ring form for 4-Oxo-2-nonenal (ONE), were built, while malondialdehyde (MDA) was directly attached to each residue. The protocol describes details for the construction, geometry optimization, assignment of charges, missing bonds, angles, dihedral angles parameters, and its validation for each modified residue structure. As a result, structural effects induced by the carbonylation with these lipid derivatives have been measured by molecular dynamics simulations on different protein systems such as the thioredoxin enzyme, bovine serum albumin and the membrane Zu-5-ankyrin domain employing root-mean-square deviation (RMSD), root mean square fluctuation (RMSF), structural secondary prediction (DSSP) and the solvent-accessible surface area analysis (SASA), among others.

Introduction

In the constant pursuit of understanding the molecular behavior of proteins with oxidative modifications, computational chemistry has become a fundamental pillar in the broad field of scientific research. This relies on the use of theoretical models capable of interpreting physical phenomena in electronic systems, using mathematical equations to describe the atomic behavior of molecules. Within this landscape, computational simulations of proteins stand out as crucial tools to analyze the atomic behavior of molecular systems. Based on the evaluation of structural behavior, energetic calculations, and conformational states¹, these methods become....

Protocol

1. Design and optimization of the new modified amino acid

NOTE: This stage involves drawing the structures of modified residues and optimizing their energy.

1. Designing the modified structures and optimizing their structure.
   1. Use a computational chemistry software package to draw the amino acid molecules bound to the reactive aldehydes derived from lipid peroxidation, i.e. with HNE, HHE, MDA and ONE. Once modified, at the carboxyl group end of the amino acid draw the shape of the methylamine group. At the amino end, draw an acetyl group to emulate the peptide bonds of the modified amino acid, as shown i....

Results

To illustrate the implementation of the protocol and evaluate the results, the following analyses will be considered. The data set generated by assigning new parameters to modified amino acid residues was constructed based on optimization of the electronic structures, which were supported for partial RESP loadings. Figure 9 shows the structural conformation of one of the amino acid residues optimized with the parameter assignment.

Discussion

One of the critical steps in developing the AMBER parameterization protocol was the quantum optimization of the new amino acid residues modified with the lipid peroxidation derivatives, due to the energetic variability related to the minimization and the way of assigning RESP charges in the AMBER antechamber. For this, ab initio optimization methods with Hartree-Fock (HF/6-31G) and semiempirical density functional theory (DFT; B3LYP/6-31G and M062X/6-31G) were established to evaluate the response to the load ass.......

Materials

Name	Company	Catalog Number	Comments
AmberTools16 or Upper	The Amber Project		Amber is a suite of biomolecular simulation programs
Gaussian 09 or Upper	Gaussian Inc		Draw and optimize structures
Linux Ubuntu	GNU/Linux		Platform for AmberTools
NVIDIA GPUs GTX 1080 or Upper	Nvidia		Compatible with PMEMD