Lecutre 13 Flashcards
What is an example of a protein that has a gpi anchor
What disease does it contribute to
The prion protein on its c terminal domain
Mad cow disease
What causes mad cow disease
Being prion protien being resistant to Proteinase K
Can’t be chopped up and causes other protiens to misfold
What is does a normal prion protein look like (prpc)
Has a hydrophobic region
Binds cu2+ in square pyramidal geometry (binds nitrogen’s so intermediate)
Has glycosylated recptors
Has an octa repeat peptide
What is the sequence of the octa repeat residues in the prion protien
PH GGG WGQ
these repeats bind the copper ions
What is the disease version of the prion protein
Prpsc
What are the types of prion diseases
Creutzfeild Jakob disease : due to old age or exposure to infected cow/human brain)
BSE: when cows are exposed to infected cow brain (mad cow diease)
Kuru: humans ingest infected human brain
Fatal familial insomnia : rare Mutation in PRNP (prion protien) gene
What are prion protiens important for
Neuroprotection
Signalling in the brain due to gpi anchor
What is the role of the phospholipase enzyme (PI-PLC)
It can cleave the gpi anchor chain from the membrane
So I release the proton and the anchor in the extracellular space
Cysteine Thiol groups are
Redox sensitive (oxidative vs reducing environment)
What is HSA
Human serum albumin
Most important protein in the blood serum
Has 7-9 fatty acid binding sites
Heart shaped
Where is HSA mainly made, as what type of protein
In the liver
As a preproprotein
Molecular weight of HSA
66kDa
The half-life of HSA is
Long
HSA in the plasma is
Very densely packed
Is HSA a glycoprotien
No
Rare because extracellular proteins are usually glycosylated
How much of protein in the blood serum is HSA
60%
Is there more HSA outside the blood vessels or inside
More HSA outside the blood vessel
What is the function of HSA in the blood vessels
To maintain the osmotic pressure of plasma inside the cells
What can you say if HSA is not found in the urine
This means that HSA doesn’t usually leave the blood
We don’t lose HSA, very kept inside the blood
What does HSA bind
fatty acids 7-9 in the plasma
steroid and thyroid hormones (like thyroxine)
Hemin
Metal ions like cu ni zn 2+ (intermediate metals)
What is hemin
Same as hemoglobin but instead of oxygen bound to fe , cl is bound
The chlorine turns fe2+ to 3+ (ferric)
Structure of thyroxine
Slide 9
How many binding sites does HSA have
Multiple
Ex. Binds 1 hemin and 7 fatty acids at the same time
HSA is mainly ______
How many domains is HSA
how many subdomains
What is it cross linked by
How many drug binding sites
Alpha helical
3
2 in each of the 3 domains
Disulphide bonds
2
Why does HSA have disulphide bonds
Because it is outside the cell in oxidize environment
How many free cysteine in HSA
1 rest are disulphide bonds
Where do the disulphide linkages occur in HSA
Only in between the subdomain in each of the three domain
None in between the three domains
Where can we make recombinant HSA and why
Only in yeast strains since ecoli can’t make disulphide bonds
Where can we make recombinant HSA and why
Only in yeast strains since ecoli can’t make disulphide bonds
What is the ATCUN site
This is the site in the HSA that binds copper and nickel at the amino terminal
Binds cu2+ and ni2+ (square planar, intermediate)
What is the most importantly amino acid in the HSA ATCUN site
The histidine
Since it’s imidazole ring binds the intermediate metals via nitrogen
What where does HSA deliver cu2+
Important do cu2+ going To the liver
What is the largest pool of copper?
Second largest
Ceruloplasmin
HSA
HSA also has a _____ binding site
Zinc2+
But doesn’t binds to the ATCUN site
What amino acids and in which domains make the zinc binding site in HSA
Between domains 1 and 2
two his side chains
What releases the zinc from the zinc binding site
If a fatty acid binds to HSA
Then the two his that bind the zinc are too far apart and the zinc is released
So zinc being there depends on fatty acid binding
How many fatty acids can bind to HSA
What types does it bind to
7-9 sites for each fatty acid
They are buried in the HSA because they’re hydrophobic
Binds both saturated and unsaturated fatty acid
What drug binds to site 1 of HSA
Site 2
Warfarin (blood thinner, prevent blood clot forming, block rectivation of vitamin K)
Diazepam (also called Valium, anti anxiety drug)
Ka=1/kd
Okay slide 17
Why does HSA bind many drugs and synthetic drugs
It’s like a drug and fatty acid buffer
If you have a drug that has a high response rate to the dose
HSA binding allows for the drug to diffuse and transport through the blood so we don’t get high dose response rate and high dose in one area
What is propofol
It’s a drug that is very dangerous because it has a high dose response rate and doesn’t bind to HSA
Killed Micheal Jackson
Where is the third drug binding site in HSA
What binds there
The 1B domain
Billiverdin and hemin bind here
Is there competition for drugs at the different binding sites of HSA
What types of interactions do these binding sites have
Yes
Allosteric interaction (binds one then the other with higher affinity)
Where is drug binding site 1 located
What about 2
Domain 2a
Domain 3a
If a drug is hydrophobic it’ll bind to
Fatty acid binding sites
What is methotraxate
How can it get displaced and what’s the result
Used as a drug in treating cancer in high doses and rheumatoid artheritis in low doses
NSAIDS (naproxen) displace methotrexate and increase free methotrexate levels in the plasma (toxic competition)
How can cancer tumours benefit with HSA
How do we stop this
They bind HSA and use it as a source of nitrogen
We can bind anti cancer drugs to HSA so the the tumour attached to HSA dies
Not specific tho, just randomly binding hsa
What another example of coupling drug to HSA
The one free cysteine (cys34) get drugs attached to it
How many cys in hsa
How many disulfide binds
What does this mean
34
17
One free cys (cys 34)
