Multifunctionality of Acidulated Serum Albumin on Inhibiting Zn²⁺-Mediated Amyloid β-Protein Fibrillogenesis and Cytotoxicity.

Fibrillogenesis of amyloid β-proteins (Aβ) mediated by transition-metal ions such as Zn(2+) in neuronal cells plays a causative role in Alzheimer's disease. Hence, it is highly desired to design multifunctional agents capable of inhibiting Aβ aggregation and modulating metal-Aβ species. In this study, we fabricated acidulated human serum albumin (A-HSA) as a multifunctional agent for binding Zn(2+) and modulating Zn(2+)-mediated Aβ fibrillogenesis and cytotoxicity. On average, 19.5 diglycolic anhydrides were modified onto the surface of human serum albumin (HSA). It was confirmed that A-HSA kept the stability and biocompatibility of native HSA. Moreover, it could inhibit Aβ42 fibrillogenesis and change the pathway of Zn(2+)-mediated Aβ42 aggregation, as demonstrated by extensive biophysical assays. In addition, upon incubation with A-HSA, the cytotoxicity presented by Zn(2+)-Aβ42 aggregates was significantly mitigated in living cells. The results showed that A-HSA had much stronger inhibitory effect on Zn(2+)-mediated Aβ42 fibrillogenesis and cytotoxicity than equimolar HSA. Isothermal titration calorimetry and stopped-flow fluorescence measurements were then performed to investigate the working mechanism of A-HSA. The studies showed that the A-HSA surface, with more negative charges, not only had stronger affinity for Zn(2+) but also might decrease the binding affinity of Aβ42 for Zn(2+). Moreover, hydrophobic binding and electrostatic repulsion could work simultaneously on the bound Aβ42 on the A-HSA surface. As a result, Aβ42 conformations could be stretched, which avoided the formation of toxic Zn(2+)-Aβ42 aggregates. The research thus revealed that A-HSA is a multifunctional agent capable of altering the pathway of Zn(2+)-mediated Aβ42 aggregation and greatly mitigating the amyloid cytotoxicity.