Unraveling the Molecular Dynamics of Glucose Oxidase Desorption Induced by Argon Cluster Collision

Abstract

The bombardment of a protein multilayer target by an energetic argon cluster ion beam enables protein transfer onto a collector in the vacuum while preserving their bioactivity (iBEAM method). In parallel to this new soft-landing variant, protein transfer in the gas phase is a prerequisite for their characterization by mass spectrometry. The successful transfer of bioactive lysozymes (14 kDa) by cluster-induced soft landing and its mechanistic explanation by molecular dynamics (MD) simulations have sparked an important inquiry: Can heavier biomolecules be desorbed while maintaining their tridimensional structure and hence their bioactivity? To address this question, we employed MD simulations using a reactive force field (ReaxFF). Specifically, the Ar cluster-induced desorption of glucose oxidase from either a gold substrate or a lysozyme underlayer was modeled using the LAMMPS code. First, the force field parameters were trained by computing the dissociation energetics of a series of organic molecules with ReaxFF and DFT, in order to realistically describe N–S and O–S interactions in the bombarded glucose oxidase molecule. Second, bombardment simulations investigated the effects of cluster size (ranging from 1000 to 10000 Ar atoms) and kinetic energy (1.5 and 3.0 eV/atom) on the structural features and energetics of the desorbing glucose oxidase. Our results show that large argon clusters (≥7000) are needed to desorb glucose oxidase from a gold surface, yet protein fragmentation and/or pronounced denaturation occur. However, the transfer of structurally preserved glucose oxidase in the gas phase is predicted by the simulations when an organic layer is used as a substrate.

Molecular Dynamics Simulations of Soft and Reactive Landing of Proteins Desorbed by Argon Cluster Bombardment

“…Our computational studies reasonably predict the entire process of molecular transfer and redeposition using large cluster ion beams, from desorption to landing of lysozymes and their noncovalent complexes, suggesting that the method could be successful with larger biomolecules . Interestingly, it also predicts a mechanism by which such hyperthermal velocity proteins can covalently graft on a substrate with minimal damage, a feature that might have significant relevance in technological applications, e.g., for biosensors with improved performance …”

“…Compared to sample sputtering by isoenergetic atomic and small cluster ions, protein desorption through iBEAM is characterized by inducing minimal damage to the molecules and has showcased its ability to transfer bioactive lysozymes (14 kDa) from a source (target) to a recipient surface (collector) . Moreover, it has demonstrated precision in constructing multilayered structures at the nanoscale, while computational simulations indicated promising prospects for the successful transfer of even larger proteins, such as glucose oxidase (64 kDa) . Notably, the extent of protein damage during desorption depends significantly on the kinetic energy per atom ( E / n ) of the GCIB, with considerable reduction in internal energy (thereby fragmentation) observed for E / n values smaller than 2 eV/atom. , However, our simulations also accentuated the role of cluster size (i.e., the number of nuclei n ) as a secondary factor in exacerbating fragmentation at constant E / n .…”

“…Recently, ReaxFF simulations focusing on the desorption of lysozymes (Lyz) and glucose oxidase (GOx) from a gold substrate were conducted in our group, revealing the potential for intact protein desorption and the dependence of fragmentation on the bombardment conditions. , An important aspect of those investigations was to shed light on the difference in molecular desorption from gold, producing proteins with high internal energies, prone to fragmentation, and desorption from a protein matrix, leading to the emission of (clusters of) proteins with comparatively lower internal energies, even with minimal denaturation. Incidentally, our simulations highlighted intriguing aspects of reactivity, such as the formation of Au–S and H–S bonds attributed to the presence of cysteine amino acids, mirroring experimental observations. , The present study capitalizes on these results to delve into the investigation of lysozyme soft and reactive landing, examining diverse scattering angles and protein configurations obtained from our previous desorption simulations in order to elucidate the influence of these variables on the landing process.…”

Gas Cluster Ion Beam-Assisted Deposition for Fabricating Dry Multilayers Containing Enzyme and Substrate with On-Demand Release

“…An absorbance measurement and a ToF–SIMS depth profile will be performed first on a bilayer GOx/β- d -glucose and then on a trilayer HRP/GOx/β- d -glucose. The choice of these proteins was motivated by MD simulations that predicted the intact desorption of GOx from an organic film …”

“…The choice of these proteins was motivated by MD simulations that predicted the intact desorption of GOx from an organic film. 21…”