Net Charge Affects Morphology and Visual Properties of Ovalbumin Aggregates

Abstract

The effect of ovalbumin net charge on aggregate morphology and visual properties was investigated using chromatography, electrophoresis, electron microscopy, and turbidity measurements. A range of differently charged ovalbumin variants (net charge ranging from -1 to -26 at pH 7) was produced using chemical engineering. With increasing net charge, the degree of branching and flexibility of the aggregates decreased. The turbidity of the solutions reflected the aggregate morphology that was observed with transmission electron microscopy. Increasing the stiffness of the aggregates transformed the solutions from turbid to transparent. Artificially shielding the introduced net charge by introducing salt in the solution resulted in an aggregate morphology that was similar to that for low-net-charge variants. The morphology of heat-induced aggregates and the visual appearance of the solutions were significantly affected by net charge. We also found that the morphology of ovalbumin aggregates can be rapidly probed by high-throughput turbidity experiments.

The Mechanism of Fibril Formation of a Non-inhibitory Serpin Ovalbumin Revealed by the Identification of Amyloidogenic Core Regions

“…In the absence of salt, OVA conformation exhibits a nearly reversible two-state heat-induced transition with a midpoint temperature of 76°C and reaches an almost completely unfolded state with a significant degree of secondary structure at 80°C (23). In the presence of salt, OVA undergoes irreversible heat denaturation with the formation of semiflexible fibrillar types of aggregates (23)(24)(25)(26)(27)(28). Unlike the other serpins, the aggregation mechanism of OVA has been suggested to resemble that of amyloid fibril formation (25,27).…”

How to tailor heat-induced whey protein/κ-casein complexes as a means to investigate the acid gelation of milk—a review

“…As a rule, environmental conditions that promote exposure of local poles of high net charges and/or β-sheet structure also promotes head-to-tail aggregation, hence fibrillation. Hence, various studies have reported that globular proteins such as ovalbumin (Weijers et al 2008), β-LG (Bolder et al 2007;Durand et al 2002;Ikeda and Morris 2002;Jung et al 2008), BSA (Holm et al 2007) or partially hydrolysed α-LA (Otte et al 2005) can yield fibrils on (even moderate) heat treatment at low ionic strength values and at pH generally~7.0 or~2.0 depending on proteins. Elongated particles of β-LG can further be produced in water at pH~7.0 and ≤50 mmol.L −1 NaCl (Alting et al 2004;Pouzot et al 2005).…”

Application Potential of Food Protein Modification

“…Cryo-EM investigation of aggregates formed from ovalbumin which had been succinylated to various degrees provided further evidence that net charge dominantly determines aggregate morphology (Weijers et al, 2008). The propensity of proteins to aggregate, or rate of aggregation, has been shown to vary as a function of protein conformational stability (Chiti et al, 2000;Hurle et al, 1994;Kelly, 1998;Quintas et al, 1997;Ramirez-Alvarado et al, 2000;Siepen & Westhead, 2002), rate of unfolding (Broersen et al, 2007a), net charge (Calamai et al, 2003;DuBay et al, 2004), and secondary structure propensity (Fernandez-Escamilla et al, 2004).…”

“…These hypotheses have been substantiated by a range of observations which involved charge introduction, removal or reversal through succinylation and methylation reactions (Broersen et al, 2007a;Weijers et al, 2008). The reactions of succinylation and acetylation both lead to blockage of the reactive amino groups of proteins with an acyl residue and are hence collectively termed acylation reactions.…”

“…Other studies show an increased emulsifying activity and stability upon succinylation of soy protein (Franzen & Kinsella, 1976a), and oat protein isolate (Mirmoghatadaie et al, 2009). In terms of gelation, an increase in net charge lead to more transparent gels upon gelation of ovalbumin which is related to the morphology of the aggregated network making up the gel structure (Weijers et al, 2008). Overall, many and detailed efforts have been made employing net charge modification of proteins in the field of food science.…”