Effects of low temperatures on the conformation of apo-lactoferrin and its interactions with α-lactalbumin and β-lactoglobulin: Application of in silico approaches

Lactoferrin (LF) is a multi-functional protein that is primarily found in milk. The apo form of LF (apo-LF) is devoid of iron, and as a result, may serve as an iron scavenger with attendant anti-pathogenic properties, thereby, endowing apo-LF with promising health benefits. Despite these advantages, the structural stability of apo-LF may be perturbed by various processing conditions. One of the strategies employed to stabilise the activity of apo-LF is to encapsulate it as part of an oligomeric complex, composed of apo-LF, α-lactalbumin (α-La) and β-lactoglobulin (β-Lg). Accordingly, the aim of this study is to investigate the structural changes of apo-LF and its interactions with α-La and β-Lg under the temperature conditions of freeze-drying (−30, 0, and 20 °C). Using in silico approaches, molecular dynamics simulations showed apo-LF had limited structural perturbation when processed at these temperatures. Both α-La and β-Lg stabilise apo-LF by enhancing and maintaining the structural integrity and cohesion of the tertiary structure of apo-LF. These stabilities are the result of electrostatic interactions, binding apo-LF and the protective proteins into a multimeric complex. The secondary structure and the anti-bacterial derivatives in apo-LF were also sustained under these simulated conditions. Furthermore, inter-residue contact maps in apo-LF predict an increase in intramolecular contacts at simulated freeze-drying conditions, resulting in agglomeration and, therefore potential proteins dispersibility during subsequent rehydration. Overall, this work demonstrates the advantages of computational approaches to understand the macromolecular conformations, interactions, and functional implications of a multimeric complex composed of apo-LF, α-La, and β-Lg at low temperatures.