Parallel β-Sheet Fibril and Antiparallel β-Sheet Oligomer: New Insights into Amyloid Formation of Hen Egg White Lysozyme under Heat and Acidic Condition from FTIR Spectroscopy

Abstract

Hen egg white lysozyme (HEWL) is widely used as a model protein for amyloid research. In this study, we aim to use Fourier transform infrared (FTIR) spectroscopy to gain new structural insights into amyloid formation of HEWL under heat and acidic condition. We reveal that the fibril-forming solution of HEWL has the capability to form fibril and oligomer with distinct β-sheet configurations under different temperatures. Amyloid fibril with parallel β-sheet configuration is formed at elevated temperature, while oligomer with antiparallel β-sheet configuration is formed at room temperature. The interplay between fibrillation and oligomerization suggests that the two β-sheet aggregates consume the same amyloidogenic materials such as peptide fragments and nicked HEWL due to lysozyme hydrolysis under heat and acidic condition. Temperature-dependent FTIR reveals that the oligomer is unstable at elevated temperature, demonstrating its off-pathway nature. The temperature-dependent formation of parallel and antiparallel β-sheet configurations discovered in lysozyme system is compared with that of amyloid-β and α-synuclein systems and the implication is discussed.

New Insight into Amyloid Fibril Formation of Hen Egg White Lysozyme Using a Two-Step Temperature-Dependent FTIR Approach

“…This is as expected. 33 On the other hand, an on-pathway oligomeric intermediate could also form at the same temperature according to our prediction. This means that we would have two different aggregates coexisting in the incubation solution.…”

“…Details of the making of the IR cell have been described previously. 33 The absorption spectrum of solvent D 2 O (pD = 2) can be taken in the same manner. Solvent subtraction from each original protein absorption spectrum is not necessary as we directly perform spectral subtraction between two original spectra to obtain difference spectrum in this study.…”

Virtual‐system‐coupled adaptive umbrella sampling to compute free‐energy landscape for flexible molecular docking

“…An experiment of hen egg white lysozyme has revealed that non-b-sheet oligomers exist as off-pathway aggregates. [37] Figure 9 indicates that the structural contributors for the shoulder (k55:5 Å ) of P strð19Þ cano ðk; v; TÞ in Figure 8 are the antiparallel b-sheets, parallel b-sheets, and the cross-structures. Note that P strð19Þ cano ðk; v; TÞ is the equilibrated distribution for the two-chain system.…”

Influence of genetic variability and external regulating factors on amyloid-beta peptide aggregation

“…Later, the research team of Eisenberg revealed the structure of an off-pathway, cylindrical oligomer of a segment of the amyloid-forming protein αB crystallin, that resembled a β-barrel composed of six antiparallel β-strands (72). We and other groups demonstrated that amyloidogenic proteins, such as Aβ, pass through an antiparallel β-sheet structured state, corresponding to oligomers, before undergoing a transition in structure to parallel β-sheet fibrils (58)(59)(60)(73)(74)(75). Recently, the group of Tycko demonstrated that Aβ , containing the FAD-linked Iowa D23N mutation and causing CAA, assembled into antiparallel β-sheet structured fibrils that were thermodynamically metastable and had a ribbon-like appearance (39).…”

“…Therapy consists mainly of treating cognitive decline and behavioural symptoms. Five cholinesterase inhibitors and one N-methyl-D-aspartate (NMDA) receptor antagonist (memantine) have been approved by the US Food and Drug Administration (FDA) for treatment of cognitive decline (75). Acetylcholinesterase inhibitors are thought to improve functioning of the cholinergic system in AD patients by binding to acetylcholinesterase in the synaptic cleft, preventing it to break down acetylcholine, a neurotransmitter involved in learning and memory (76).…”