Vl 1 - metabolic pathways Flashcards
what are TgFBP2 and TgPFK used for?
TgFBP2: Fructose-1,6-bisphosphate ⇒ Fructose-6-phosphate + Pi ⇒ glycan synthesis (storage)
TgPFK: Fructose-6-phosphate + Pi ⇒ Fructose-1,6-bisphosphate ⇒ gylcolysis
- appear to form cycle which may represent a novel form of metabolic regulation
- use one or the other, depending on what is needed
- Utilization of glucose for glycan synthesis
- Phosphate homeostasis during fluctuating glycolytic flux.
TgFBP1 and TgFBP2
Fructose 1,6,bisphosphatase 1/2
Fructose-1,6-bisphosphate ⇒ fructose-6-phosphate + Pi
- TgFBP2 does most work - essential during glucose metabolism (controls hexose phosphate dependant pathway) ⇒ required for parasite growth
- TgFBP2 knock down increases glycolytic flux and inhibits the TCA cycle and FAS2
- also interferes with glycan synthesis
- dysregulates steady state levels of lipid glycans and dolichol lipids
CDPK2
calcium dependent protein kinase 2
- binds and controls amylopectin turnover
- Important for the development of Toxoplasma Bradyzoites ⇒ good drug target
how can we measure the metabolome?
with high resolution by mass spectrometry
Discoveries led by metabolome
- GABA-shunt
- mediates both the catabolism of GABA and channels L-glutamate to the TCA cycle III (animals), bypassing two steps of that cycle
- GABA is made from glucose and glutamine in intracellular parasites (acts as energy source)
what do Antifolates do?(are Block buster drugs)
- block folic acid (vit. B9)
- Pyrimethamin: blocks dihydrofolat reductase (needed for activating folic acid)
Sulfadoxin: competetively blocks dihydropteroate synthase (found in bacteria, plants and fungi) ⇒ inhibits folic acid synthesis
1) which is the alternative to mevalonate pathway in apixomplexans?
2) how is the mevalonate pathway also called?
3) which metabolites are needed for IPP and DMAPP synthesis in mamals and apicomplexans?
1) Non-Mevalonat-Pathway, (MEP-/DOXP-Pathway)
2) Isoprenoid-Pathway.
3) mammals: Acetoacetyl-CoA and Acetyl-CoA;
Apicomplexans: pyruvate and G3P.
describe the advantages of the non-mevalonat metabolism in apicomplexans for drug development
- mammals: synthesize DMAPP and IPP via mevalonat pathway in the cytosol
- parasites use MEP-/DOXP-pathway (Non-Mevalonat-Pathway)
- different DMAPP synthesis pathways in parasite and mammals ⇒ develop drugs that only targed parasite specific parts
- e.g.: fosmidomycin (inhibits DXP Reductoisomerase ⇒ key enzyme in parasite MEP-/DOXP-pathway) inhibits DOXP ⇒ MEP
why are DMAPP and IPP vital?
important for:
dolichol Lipids, glycolysation, protein prenylation, Ubiquinone, Q10, Electron Transport Chain, Isopentylation of t-RNA.
- primary reactant for isoprene biosynthesis (DMAPP)
- cellular terpenoid biosynthesis (both)
why does plasmodium possess an Apicoplast?
- location of IPP (and DMAPP) synthesis
- IPP and DMAPP play important role in many metabolic pathways
(nedded for production of Terpenes and Terpenoids). - all these pathways are inhibited, if assembly of apicoplast is impaired.
ENDOSYMBIOSE?
how to inhibit IPP synthesis in plasmodium?
With Doxycyclin:
⇒ blocks binding of tRNA to mRNA ⇒ AA can’t be assembled to polypeptide chains ⇒ no new proteins
⇒ stops growth ⇒ immune system can kill parasites
how to make a flux-balance model?
1) examine gene expression
2) identify biomass composition (lipids, polysaccharides, proteins, vitamins, nuclelic acids)
3) reconstruct metabolic network
4) draft restrictions
5) calculate optimal flux distribution
what are dispensable metabolites? exsample
- spezific metabolite molecule not important, can be exchanged by another
- e.g.: needs one of the sugars - doesn’t matter which one (D-Fructose, D-Glucose, D-Mannose, etc),
- source of anorganic iron: heme or Fe2+.
- opposite is indispensable ⇒ molecule can’t be exchanged by another
- e.g.: Cholesterol, L-Arginin.
can T.gondii synthesize glucose by itself? Why is it needed? how is it stored?
- no, uses host sugars for energy and synthesis of glycoconjugates (important for survival and virulence)
- intake via TgGT1 (4 different glucose transporters - only TgGT1 active)
- sugar stored as amylopectin
- glutamine can serve as energy source
glutamine supports virulence, when glucose is absent
Stable isotope-resolved dissection of parasites by GC/MS. what does it tell us? how does it work? how do you work with it?
- estimates relative frequency and turnover ratio of metabolites (simultanious production and usage)
- stable isotopes e.g.: 2H/13C (dont degrade, not radioactive) needed for turnover ratio measurement
- using heavy U 13C6-glucose (all carbon atoms 13C, more mass) ⇒ all heavy glucose derived molecules have higher mass
- e.g.: glucose-6-phosphate has 6 g/mol more
- citric acid has 2,4 or 6 g/mol more, depending on repeatedly integrated acetyl-coa
⇒ change in mass can be used to determine active pathways and measure metabolite fluctuation ratios
