Effects of arginine on heat‐induced aggregation of concentrated protein solutions

Abstract

Arginine is one of the commonly used additives to enhance refolding yield of proteins, to suppress aggregation of proteins, and to increase solubility of proteins, and yet the molecular interactions that contribute to the role of arginine are unclear. Here, we present experiments, using bovine serum albumin (BSA), lysozyme (LYZ), and β-lactoglobulin (BLG) as model proteins, to show that arginine can enhance heat-induced aggregation of concentrated protein solutions, contrary to the conventional belief that arginine is a universal suppressor of aggregation. Results show that the enhancement in aggregation is caused only for BSA and BLG, but not for LYZ, indicating that arginine's preferential interactions with certain residues over others could determine the effect of the additive on aggregation. We use this previously unrecognized behavior of arginine, in combination with density functional theory calculations, to identify the molecular-level interactions of arginine with various residues that determine arginine's role as an enhancer or suppressor of aggregation of proteins. The experimental and computational results suggest that the guanidinium group of arginine promotes aggregation through the hydrogen-bond-based bridging interactions with the acidic residues of a protein, whereas the binding of the guanidinium group to aromatic residues (aggregation-prone) contributes to the stability and solubilization of the proteins. The approach, we describe here, can be used to select suitable additives to stabilize a protein solution at high concentrations based on an analysis of the amino acid content of the protein.

Inhibition of protein aggregation by zwitterionic polymer-based core-shell nanogels

“…The same trend was observed when the nanogel concentration was increased (Figure S5). In the presence of NG-F, less than 10% fibrillation is observed even after heating at high temperatures for 30 minutes, a value that is much lower than that obtained with linear polymers (poly-SPB) and previously reported compounds such as non-detergent sulfobetaines3334353637 and L-arginine hydrochloride3839 (Figure S6). …”

Small Molecules Present in the Cerebrospinal Fluid Metabolome Influence Superoxide Dismutase 1 Aggregation

“…Indeed, many of the ALS pathological mutations are suggested to lead to increased aggregation propensity due to these effects [54]. In addition, arginine which is frequently considered as a beneficial aggregation suppressor by enhancing the solubility of proteins [55,56], actually promoted apoSOD1 aggregation, an effect also evidenced in other proteins such as bovine serum albumin and β-lactoglobulin [57]. The versatility of arginine to suppress/promote protein aggregation is thought to rely on the preferential binding of its methyl guanidinium group, which may either interact with aromatic side chains (suppressing aggregation) or with acidic side chains (promoting aggregation).…”

Inhibitory Effects of Arginine on the Aggregation of Bovine Insulin

“…While the mechanism of arginine's interaction with insulin and with other proteins is not yet well understood [32–36, 41, 42] it appears to be related to its binding to aromatic groups in partially unfolded states of proteins and may therefore be an effective inhibitory agent for a wide variety of amyloid forming proteins.…”

“…The role of the guanidinium group of arginine binding to aromatic groups of insulin when these are exposed in misfolded monomers leading to inhibition of aggregation has been suggested by Lyutova et al [35]. A recent paper by Shah et al [36] reports that arginine can also act to promote aggregation of proteins and that the key as to whether it inhibits or promotes aggregation appears to be whether the guanidinium group binds to aromatic or acidic residues, respectively. …”