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Sources of Amino Acids De Novo Synthesis CO 2 fixation (ala, asp, glu) little incorporated into protein Host Plasma uptake of all amino acids in vitro.

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Presentation on theme: "Sources of Amino Acids De Novo Synthesis CO 2 fixation (ala, asp, glu) little incorporated into protein Host Plasma uptake of all amino acids in vitro."— Presentation transcript:

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2 Sources of Amino Acids De Novo Synthesis CO 2 fixation (ala, asp, glu) little incorporated into protein Host Plasma uptake of all amino acids in vitro growth requires ile, met, cys, gln, glu Digestion of Host Hemoglobin

3 Hemoglobin 95% of total erythrocyte protein very abundant (>300 mg/ml or approximately 5 mM) 60-80% is degraded during erythrocytic stage 110 g (of 750 total) is consumed in 48 hrs at 20% parasitemia

4 Endocytosis of Host Cytoplasm food vacuole cytostome pinocytosis (rings)

5 The Food Vacuole A Specialized Lysosome ATP ADP H+H+ (pH 5-5.4) Food Vacuole Proteases plasmepsins I & II (acid) falcipains I - III (thiol) falcilysin (metallo) Absent: other acid hydrolases hemoglobin digestion parasite cytoplasm Endocytic Pathway

6 Proteases Mediate the Catabolism of Proteins proteases (aka peptidases) break the peptide bonds that hold amino acids together exopeptidases remove amino acids sequentially from either N- or C-terminus endopeptidases cleave between specific residues within polypeptide chain

7 Initial plasmepsin cleavage is specific and leads to a destabilization of hemoglobin native Hb is cleaved between Phe-33 and Leu-34 ( chains) –hinge region –conserved –important for tetramer stability the large globin fragments dissociate –heme is released globin fragments are susceptible to further proteolysis F33/L34

8 Hemoglobin Digestion is an Ordered Process exopeptidase? free amino acids? hemoglobin + heme large globin fragments small fragments (6-8 amino acids) plasmepsin falcipain plasmepsin medium fragments (20 amino acids) falcilysin

9 ABC Transporter Super Family large and ubiquitous gene family defined by ATP-Binding Cassette aka Multi-Drug Resistance (MDR) transport is usually specific for particular types of substrates Pfmdr-1 protein localized to food vacuole Pfmdr-1 complements yeast ste6 gene ste6 transports a-type mating factor (12 residue peptide)

10 The Food Vacuole A Specialized Lysosome hemoglobin +heme globin fragments small fragments (6-8 amino acids) ATP ADP ATP ADP H+H+ plasmepsin falcipain plasmepsin falcilysin Pfmdr-1? amino acids amino- peptidase ABC transporter associated with food vacuole amino-peptidase activities in parasite cytoplasm

11 heme destabilizes and lyses membranes hydrolases released into parasite cytoplasm parasite dies Free Heme is Toxic Possible Detoxification Mechanisms heme hemazoin (malaria pigment) H 2 O 2 mediated degradation GSH mediated degradation heme oxygenase (P.b. and P.k. only)

12 Hemazoin = -Hematin heme -hematin

13 -hematin forms insoluble crystals 'biocrystallization' or 'biomineralization'

14 Pigment Formation biocrystallization mechanism unknown beta-hematin can form spontaneously (harsh conditions) histidine-rich proteins or lipids can promote the process heme biocrystallization inhibited by chloroquine and other anti-malarials

15 The Food Vacuole A Specialized Lysosome hemoglobin + heme globin fragments small fragments (6-8 amino acids) hemazoin ATP ADP ATP ADP H+H+ Fe 3+ Fe 2+ O2O2 - O 2 O 2 ? plasmepsin falcipain plasmepsin falcilysin Pfmdr-1? amino acids amino- peptidase iron oxidized after release from Hb oxidation promotes formation of ROI oxidative stress

16 The Food Vacuole A Specialized Lysosome hemoglobin + heme globin fragments small fragments (6-8 amino acids) hemazoin ATP ADP ATP ADP H+H+ Fe 3+ Fe 2+ O2O2 H2O2H2O2 H 2 O + O 2 - O 2 O 2 superoxide dismutase? catalase? ? plasmepsin falcipain plasmepsin falcilysin Pfmdr-1? amino acids amino- peptidase


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