Lecture 20 - New drug targets for malaria Flashcards
Describe the vacuolar membrane
When the parasite is invading an RBC, it pushes through the membrane, which ends up surrounding it inside the RBC
Parasitophorus vacuole allows the parasite to develop while protected from the phagolysosomes of the host cell
What is KAHRP? What happens during KO?
One of the hundreds of exported proteins important for virulence
First gene knockout in a malaria parasite
Essential for knob formation and adherence
* Rough appearance w/ PfEMP1 present
* The knobbly rough structures are what PfEMP1 is embedded into
* Allow the parasite to hang onto to the endothelium under physiological flow pressure
* KHARP is essential to these knobbly structure
KnockOut:
* smooth appearance
* can’t adhere to vascular endothelium
* PfEMP1 is still there, but it can’t hang on to the endothelial walls under physiological flow pressure
What are the ‘Hundreds of proteins’?
Give an example of one
Hundreds of proteins (500) that are exported out of the parasite to
perform a specific function.
* It doesn’t matter for us what this function is, but we will
look at how they are transported out of the parasite
* Crucial to the parasite (virulence or life cycle)
* This could be a good target for a drug
KAHRP
* Exported protein vital for rough appearance of the infected RBC
* Crucial for infected RBC adherence to the endothelium
What is the PEXEL motif?
Why is it important?
Plasmodium export element
* Of the exported proteins that were known, all had a common looking N-terminus.
It had:
* a hydrophobic region (signal sequence
* as well as a motif: PEXEL
Motif:
RxLx(E/Q/D)
Where ‘x’ is any amino acid
Importance:
* This motif is responsible for export out the protein out of the vacuolar membrane
- If any of the residues in the PEXEL motif, GFP was not able to be exported out of the vacuolar membrane
- The proteins with the PEXEL motif have important functions for the life cycle of the parasite:
- Cytoadherence
- Waste clearance
- Membrane rigidity
- Nutrient delivery
- Thus, they can not be KO’d
- Thus, PTEX is a good target for a malarial drug
What was seen in knockout screens of 51 exported proteins?
What is the significance of this?
PTEX is a great drug target
(46 were PEXEL containing)
* The genes have very varied functions.
* A quarter of these genes could not be knocked out
* The genes were essential to the blood stage metabolism.
* This would be a great drug target.
* A drug wants to kill a parasite
NB It is thought that almost all of these proteins have a common transport mechanism
How do the proteins get across the vacuolar membrane?
Parasite → Vacuolar space
* In the ER of the parasite
* Plasmepsin V cleaves PEXEL
RxLx(E/Q/D) → RxL + x(E/Q/D)
* Normal vesicular trafficking to the a very particular area of the plasma membrane of the parasite
* Vacuole contents released into vacuolar space
Vacuolar space → RBC cytosol
Movement through a translocon: PTEX
Proteins recognise machinery of translocon (?, still unconfirmed)
Hsp101 unfolds proteins so that they can pass through
Passes through EXP2 pore
Refolded by other subunits on the other side
What are the key criteria of the PTEX translocon?
Plasmodium specific
Essential to blood stages
Energy source, because all translocons use ATP or GTP
Must bind transiting cargo PEXEL proteins
What is the structure of the PTEX translocon?
5 protein subunits
Hsp101:
* A heat shock protein
* The energy source
* Unfolds proteins
EXP2
* Forms a pore through the plasma membrane
What is the function of the PTEX translocon?
The PTEX translocon is a protein complex that is essential for the survival of the malaria parasite Plasmodium falciparum. The PTEX translocon functions as a secretion system that is used by the parasite to export proteins from its cytoplasm to the host red blood cell. This is an important mechanism for the parasite to manipulate its host and avoid detection by the immune system.
What are the best targets for a malaria drug?
How about a vaccine?
Drug:
1. PTEX:
* It plays a vital role in the movement of hundreds of proteins out of the vacuolar space
Vaccine:
2. Invasion ligands
* EBA’s
* PfRH’s
- NB Too variable, vaccine / drug would need to address all
PfEMP1
* Too variable
Are the EBA and PfRh family of receptors alternatives for the entry of malaria into red blood cells, or are they co-receptors in addition to the PfEMP1?
PfEMP1 proteins are used for cytoadherence of infected red blood cells to the vascular endothelium and to achieve antigenic variation to avoid recognition by pre-existing antibodies. The EBA and Rh proteins are ligands involved in erythrocyte binding to allow invasion of the erythrocytes by the parasite.
Is Plasmepsin V important for export of proteins across the parasite membrane or across the parasitophorous vacuole membrane? Is Plasmepsin V considered a good drug target? or just PTEX
Plasmepsin V is an aspartyl protease that cleaves the protein at the PEXEL motif. This motif needs to be cleaved for export out of the parasitphorous vacuole membrane through PTEX. So I’d say both PTEX and Plasmepsin are potential drug targets because they would prevent the export of proteins that result in virulence and growth of the parasite.
