Protein Aggregation and Performance Optimization Based on Microconformational Changes of Aromatic Hydrophobic Regions

Abstract

Protein aggregation is a key concern in biopharmaceutical development and manufacturing. There is growing interest in understanding how the changes in protein microconformation affect the aggregation behavior. This study selected several representative proteins and first manipulated microconformational changes of the aromatic hydrophobic regions of proteins, especially tryptophan residues, by using amine or guanidine additives. The effects of the interactions between the additives and proteins on the aromatic hydrophobic regions could be grouped into three categories: exposure to solvent, burial into core, and no change. The microconformational parameters of the tryptophan residue, including fluorescence peak position (λ), degree of hydrolysis, solvent accessible surface area ( SAS), and packing density ( Den), were obtained by steady-state fluorescence spectroscopy, proteolysis coupled with electrophoresis, and molecular dynamics simulation. Furthermore, the aggregation degrees of globular proteins with distinct surface aromatic hydrophobilities under mechanical stress were investigated. A strong correlation was observed between protein aggregation and the microconformational changes of the aromatic hydrophobic regions incurred by amine or guanidine additives. Protein aggregation was enhanced when the aromatic hydrophobic regions were exposed to the solvent but suppressed when the additives led to burial of the aromatic hydrophobic regions with lower-polarity microenvironment. These findings shed light on the relationship between protein aggregation and molecular conformation and paved way for future preformulation studies of therapeutic proteins.

Hampering the early aggregation of PrP-E200K protein by charge-based inhibitors: a computational study

“…These non-covalent interactions (hydrogen bonding and charge–charge interactions), as well as the aromatic contacts, are typical of molecules able to bind amyloidogenic proteins, especially of amyloid inhibitors derived from natural products [ 73 – 75 ]. In this view, the interaction with Tyr169 may be significant by considering that it has been proposed how interactions can hamper the proteins aggregation process and protein misfolding [ 76 ], and suggested that aromatic interactions may be crucial in the aggregation process [ 77 ]. Moreover, it is worth noting that the Tyr169 is strictly conserved in mammalian PrPs likely implicated in the self-assembly process of E200K monomers through its involvement in stacking that are crucial for the formation of amyloid core [ 78 – 80 ].…”

Effect of the heating process on the physicochemical characteristics and nutritional properties of whole cotyledon soymilk and tofu

“…36 Fiber particles with large specic surface area were easily adsorbed by macromolecular protein aggregation, which result in particle size of protein-ber aggregate increasing quickly. 37 3.3. The varying tendency of the particle size distribution of whole cotyledon soymilk during the heating process…”

Inhibitory Effect of DMSO on Halohydrin Dehalogenase: Experimental and Computational Insights into the Influence of an Organic Co‐solvent on the Structural and Catalytic Properties of a Biocatalyst

“…When protein is exposed to a denaturing agent, the shift in its size to larger particles is commonly ascribed to random protein rearrangement into a looser structure and molecular aggregation [53] . Under harsh conditions, denaturation and protein unfolding may occur, followed by rapid and unspecific aggregation due to hydrophobic interactions among unfolded chains [55] . This effect occurs faster at higher concentrations of OSs, especially hydrophilic ones [31] .…”

“…[53] Under harsh conditions, denaturation and protein unfolding may occur, followed by rapid and unspecific aggregation due to hydrophobic interactions among unfolded chains. [55] This effect occurs faster at higher concentrations of OSs, especially hydrophilic ones. [31] In 40 % DMSO, after 5 h, the smallest HheC particles were about 700 nm in size (Figure 4C).…”