Amyloid Diseases 1 Flashcards
Amyloid formation & amyloidoses
Amyloid is formed by the self-assembly of soluble monomeric precursor peptides
- soluble monomers become an insoluble fibril with a cross-beta molecular architecture
Can cause amyloidoses where amyloid formation is due to a protein misfolding, causing disease:
- Parkinson’s, Alzheimer’s, Type 2 Diabetes…
Structure & properties of amyloid fibrils
Rope-like structures
- can be microns longs
- typically composed of 2+ protofilaments (subunits in protofilaments have cross-beta structure)
- Fibrils can form large assemblies (extracellular = plaques; intracellular = inclusions)
The Cross-Beta fold
Beta strands stack on top of each other into a ‘rungs-of-a-ladder-like’ arrangement perpendicular to fibril’s long axis
- depending on monomer, subunit arrangement may differ
- structurally observed using solid state NMR & cry-electron microscopy
Protofilaments assemble into amyloid fibrils
2-6 protofilaments –> 1 amyloid fibril
- protofilaments twist together to form unbranched rope-like fibrils of 30-150nm in diameter and many microns in length
Amyloid fibrils forming large assemblies
Fibrils deposit into large assemblies (plaques or inclusions) in a mesh-like network: sequestering other molecules
- extracellular plaques (e.g. Alzheimer’s)
- intracellular inclusions (e.g. Lewy bodies in Parkinson’s)
Detecting amyloid fibrils
Using dyes:
- Congo Red
- Thioflavin T
Using specific antibodies:
- WO1
Amyloidogenic precursors are either unfolded or must be partially unfolded
Forming cross-B structure requires rearrangement of peptides
Intrinsically unfolded precurors:
- a-synuclein in Parkinson’s
- B-amyloid (Ab) peptides in Alzheimer’s
Globular proteins can form amyloid but require partial unfolding/destabilisation; achieved by:
- Acidic pH, protein mutations or proteolysis producing amyloidogenic fragments
B2-microglobulin as an amyloidogenic precursor
At pH7 is very stable (no aggregation) but at pH2 (in vitro) structure is lost and readily forms fibrils
Also removing N-terminal 6 residues (by proteolytic cleavage) destabilises protein structure (even at pH7)
Association of lysozyme mutation and amyloidosis
Human Lysozyme amyloidosis is an inherited disorder
- Point mutations in the protein: Ile56Thr + Asp67His mutations destabilise lysozyme structure and enhance amyloid formation
Oligomer formation
Soluble oligomers are formed in the Lag phase: small aggregates form that are distinct from insoluble fibrils:
- can be heterogenous and short lived (hard to study)
- Can be intermediates or part of dead-end assembly pathways
- Some are toxic and may be the principle toxic agent in amyloid disease
Mechanisms of Amyloid Toxicity: Plaques
Plaques are a key feature in diseases such as Alzheimer’s
- can disrupt tissue architecture/invade extracellular space
- poor correlation between plaque deposition and neuronal disruption in Alzheimer’s (possibly a different culprit…)
Mechanisms of Amyloid toxicity: Oligomer cytotoxicity
Oligomer presence in vivo correlates better with neuronal disruption in Alzheimer’s than plaques (oligomers = cytotoxic; plaques = inert end products)
- The ‘A11’ antibody can protect against oligomer cytotoxicity
Oligomers mechanism of killing cells
1) disrupting purified lipid membranes (liposomes)
2) Disrupting plasma membrane
- inducing Ca2+ influx to promote apoptosis
these mechanisms aren’t observed when in monomeric or fibril form
Mechanisms of amyloid toxicity: fibrils sequestering other proteins
Taking away important cellular proteins has functional consequences
- e.g. Sequestering Nuclear Pore proteins, disrupts nuclear export of RNA and proteins
Fibrils = alternative source of oligomers
Depolymerisation of fibrils can produce cytotoxic oligomers
- e.g. Depolymerisation of a-synuclein fibrils produce cytotoxic oligomers
