Amyloid Fibril Formation and Seeding by Wild-Type Human Lysozyme and Its Disease-Related Mutational Variants

Detection of protein aggregation with a Bloch surface wave based sensor

“…HEWL samples are dissolved in concentrations from 35 M to 1.4 mM (0.5 mg/ml to 20 mg/ml) in a 10 −2 M HCl solution (pH 2). In order to generate lysozyme fibers, the mixtures are incubated for 1 week at 65 • C [23]. The protein concentrations are determined by absorption measurements at 280 nm before and after incubating the samples.…”

Probing the origins, diagnosis and treatment of amyloid diseases using antibodies

“…In order to carry out detailed studies of such processes in vitro, it is therefore necessary to increase considerably the rates at which they occur. For globular proteins, one way of achieving this objective is to employ conditions that favour the formation of at least partially unfolded states, for example low pH values [25], high temperatures [26], low to moderate concentrations of strong denaturants [27,28], or the presence of organic solvents [13,29]. Second, the structural characterisation of the various species formed during the process of fibril formation is complicated by their heterogeneity and by their transient or insoluble nature.…”

The Extracellular Chaperone Clusterin Potently Inhibits Human Lysozyme Amyloid Formation by Interacting with Prefibrillar Species

“…In-depth characterization of two of the disease-associated variants, I56T and D67H, as well as the non-diseaserelated T70N variant, has been performed using a variety of biophysical techniques. [7][8][9][10][11][12][13][14] Deposits of I56T and D67H formed in vitro possess the key characteristics of amyloid fibrils; these include their fibrillar morphology in TEM images, their displays of birefringence upon binding the dye Congo red and a typical cross-β X-ray diffraction pattern. [10][11][12] The kinetics of in vitro fibril formation by both I56T and D67H are sigmoidal, showing a lag phase followed by an exponential growth phase typical of amyloidogenic systems.…”

“…[7][8][9][10][11][12][13][14] Deposits of I56T and D67H formed in vitro possess the key characteristics of amyloid fibrils; these include their fibrillar morphology in TEM images, their displays of birefringence upon binding the dye Congo red and a typical cross-β X-ray diffraction pattern. [10][11][12] The kinetics of in vitro fibril formation by both I56T and D67H are sigmoidal, showing a lag phase followed by an exponential growth phase typical of amyloidogenic systems. [10][11][12] Furthermore, fibril formation by I56T, D67H and wild-type lysozymes can be accelerated by seeding with pre-formed fibrils, 12 indicating that lysozyme fibril formation proceeds through a nucleation-dependent mechanism.…”