Week 9.1 AD Flashcards
Where do Ab come from?
How are molecular details identified?
What does X-ray diffreaction indicate?
How is X-ray diffraction conducted?
What is cross-beta motif?
Ab comes from: Assemblies called amyloid fibres which are cleaved from APP
Molecular details from: EM, fibre diffraction, ss-NMR and H/D exchange
**X-ray diffraction- Indicates cross-beta motif **
Fibres that have been stretched out with a capillary tube to align, and haven’t crystalized together –very low resolution.
From this a cross-beta motif can be inferred, which are inter-molecular beta-sheet at right angles to long axis of fibre
Cross-Beta motif:
inter-molecular beta-sheet at right angles to long axis of fiber
What is fibril model of Ab baseed on?
What did it tell us about intra-molecular structure?
Fibril model of Ab: based on solid state NMR
The N terminus of Ab (1-12), loop between 2 b strands and salt bridge between the ends of the lipid: -ve charged aspartate & +ve charged lysine
Intra-molecular b-strand residues (13-22) and (29-40)
Loop with salt bridge between Asp23-Lys28
How is the stability of the fibres identified?
Stability of the fibres is identified by using Hydrogen–deuterium exchange and NMR to detect protons replaced by Du,
Hydrogen bonded = proton will not be replaced
Not hydrogen bonded = proton is replaced
**Fibril Morphology **
**What conditions affect the morphology of mature fiber **
**Are daughter fibres affected by shaking? or does the ‘seed’ determine the structure? **
In vetro-The morphology of the mature fiber is dependant on conditions
Agitation
Concentration
pH
NaCl
Robert Tycko (2013)
2 structural fibres, depending on shaking conditions: no shake/shake
The structure of the seed you add will influence morphology >
Daughter fibres – doesn’t matter if you shake or not you will end up with the same morphology
The data suggest that single fibre morphology can spread from a single nucleation site through out the brain. Structural variations may correlate with variations in AD pathology
Idea of spreading (seed) disease-concept of once you have nucleating misfolded fibre (seed) disease- that will spread to rest of brain and this will spread and have same type throughout brain.
Genetics of AD
- *What percentage of cases of AD are inheritied i.e familial? **
- Where are the main two mutations found? **
Early onset familial AD, 5% of all cases,
Inherited (early onset) familial AD
Genetic alterations underlying familial AD involve increase the production and/or the deposition of Ab in the brain. Making a strong direct link between Ab and AD. Mutations in APP or near where it is cleaved or in the enzymes that cleave it
<!--[if !supportLists]-->
• <!--[endif]-->Evidence that Ab deposition plays a pivotal role in AD comes from several lines of inquiry. Perhaps the most compelling is the genetics of inherited forms of AD.
<!--[if !supportLists]-->
• <!--[endif]-->Ab is produced by the proteolysis of a transmembrane amyloid precursor protein (APP).
<!--[if !supportLists]-->
• <!--[endif]-->Familial AD patients have been found to have mutations in APP,
OR
<!--[if !supportLists]-->
• <!--[endif]-->Within presenilin-1 (PS1) and Presenilin–2 (PS2). Presenilin proteins are key protein in the g-secretases complex - involved in the processing of APP.
- *Genetics of AD**
- *Early on set cases**
What mutation do the majority of fAD have?
How many mutations have been found in PS1? what does this cause? hint: ratio
Majority have presenilin (PS) mutations (accounts for 25% of the 5% total)
Presenilin-1 (PS1) in the g-secretases complex
More than 100 mutations identified are linked to familial AD as they increase the Ab42/40 ratio
Decides in where it will be cleaved, more cleaved at 42 which is more prone to forming fibres
What about mutations in APP itself?
How many known mutations linked with APP?
What does this mutation affect?
Mutations in APP itself
Either just before the cleavage site or just after, mutations likely to affect how these secretases cleave out Ab and the length at which it is cleaved
Up to 25 different mutations linked to APP
_Mutations occur in loop region – between the beta strands _
**Mutations in Ab sequence itself: single point mutations **
List the types:
**What do all of them apart from the Flemis affect on Ab? **
Mutations in Ab itself: Single point mutations:
Dutch type (E22Q),
Arctic (E22G),
Italian (E22K),
Iowa (D23N),
Flemish (A21G).
All (but the Flemish) mutations typically make Ab more prone (accelerated fiber kinetics) to fibril and oligomer production - form more oligomer (which are thought to be more toxic)
i.e the fibers form more rapidly - and this correlates to when you will get AD
Cluster of residues are important in triggering the early onset of AD: residues 21, 22 and 23
What affect do the mutations have?
-Loss of negative charge and the Loss of Asp23(D) and Glu22(E) and the raise pI of Ab so less soluble – this has impact on the overall charge of the protein, lowering the pH will increase fibre formation 100x’s
OR
-Addition of Gly in loop so may destabilize monomer, making fibre formation more prone to happen
Down’s Syndrome link to development of early onset AD
How many births resutls in DS?
What chromosome shows trisomy?
What do DS eventually suffer?
Where is there gene coding for APP found?
How much more likely are DS suffers likely to get AD?
<!--[if !supportLists]-->
• <!--[endif]-->Down’s syndrome occurs in 1 in 600 of births and is characterised by an extra copy of chromosome 21.
<!--[if !supportLists]-->
• <!--[endif]-->All people with Down’s syndrome develop the neuro-pathological features of Alzheimer’s.
<!--[if !supportLists]-->
• <!--[endif]-->Strikingly, a gene found on chromosome 21 encodes APP.
<!--[if !supportLists]-->
• <!--[endif]-->It is believed that the over expression of this gene alters the levels of Ab
<!--[if !supportLists]-->
• <!--[endif]-->Chromosome 21 (extra copy) and it is where APP is expressed and people with DS express twice as much APP as the rest of the population and form early onset AD.
**Genetic Risk factors in Sporadic cases of AD **
What is the main genetic factor?
Early onset AD caused by two genetic factors: ?
Depending on which allele combination main genetic factor….
Genetic Risk factors in Sporadic AD
The main genetic risk factor is having a particular form of APOPE: _APOE Apolipoprotein E (mediates cholesterol metabolism) _People with sporadic form, (25%) have apoe-e4 allele
<!--[if !supportLists]-->
• <!--[endif]-->Mutated Presenilin or APP genes cause AD for the majority of early onset AD patients (5% of cases)
<!--[if !supportLists]-->
• <!--[endif]-->Increased risk (3 fold) of contracting late onset AD is associated with APOE-e4 allele (25% of cases)
<!--[if !supportLists]-->
• <!--[endif]-->Two copies of APOE does not cause AD but increases the risk of developing it by an
8 fold increase
<!--[if !supportLists]-->
• <!--[endif]-->APOE is responsible for the transport of intracellular lipids.
<!--[if !supportLists]-->
• <!--[endif]-->ApoE e4 allele does not affect the production of Ab but does increase steady state amount
<!--[if !supportLists]-->
• <!--[endif]-->Depending on which combination of alleles i.e e4+e4 you have 7/10 chances of developing AD
Why does APOPE increase the risk?
Where does it bind?
What affects does it have?
Influences _____ homeostasis
Not exactly known
Binds to Ab and enhances plaques
Effects clearance of Ab across Blood Brain Barrier
Cholesterol homeostasis-, which in turn will affect cleavage of APP in the membrane, because cholesterol is found in lipid rafts, lipid bilayers are where APP is cleaved, and Ab is generated
GWAS studies on LATE onset revealed?
Confirms which genetic risk factors?
Risk factor for LATE onset of AD: GWAS
Genome wide associated studies, by looking at genomes of AD patients and compare with people without AD
Confirms APOE e4 as key risk factor
Clusterin, CR1 and PICALM also linked to late onset AD
- Links with: extracellular chaperone; lipid transport (APP cleavage) and Inflammation
What is the amyloid cascade hypothesis?
Missense mutations in APP, PS1, or PS2 genes
V
Increased Ab42 production and accumulation
V
Ab42 oligomerization and deposition as diffuse plaques
V
Subtle effects of Ab oligomers on synapses
V
Microglial and astrocytic activation (complement factors, cytokines etc, etc.)
V
Alterted neuronal ionic homeostasis: oxidative injury
V
Altered kinase/phosphatase activies –> Tangles of TAU
V
Widspead neuronal/neuritic dysfunction and cell death with transmitter defictis
V
Dementia
**The Tau Protein **
What is the normal function of tau?
After altered kinases and phosphatases acivity what group get phosphorylated, on which aa’s?
What affect does this have?
<!--[if !supportLists]-->
• <!--[endif]-->Tau normal function is to bind and stabilize axonal microtubules within the cell
**When the microtubules fall apart the cell loses its structure **
<!--[if !supportLists]-->
• <!--[endif]-->Altered kinases and phosphatases activity
<!--[if !supportLists]-->
• <!--[endif]-->the OH groups of Ser and Thr become phosphorylated
<!--[if !supportLists]-->
• <!--[endif]-->Tau protein is phosphorylated this causes Tau to become disassociated from the microtubules (cannot bind to microt. and is released).
<!--[if !supportLists]-->
• <!--[endif]-->On surface of microtubules
<!--[if !supportLists]-->
• <!--[endif]-->Active phosphorylation means Tau cannot bind to microtubules and its released.
