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Biochemistry of Plasmodium
Mark F. Wiser
http://www.tulane.edu/~wiser/malaria/fv.html
MACROMOLECULE
DNA
RNA
BUILDING MAJOR
BLOCK
FUNCTION
Nucleotides
Genetic Material
Template for
Protein synthesis
Proteins
Amino
acids
Cell Structure and
Function
Lipid
Fatty
acids
Membrane
Component
Carbohydrates
Sugars
Energy
Production
Sources of Amino Acids
 De Novo Synthesis
 CO2 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
Hemoglobin
 95% of total erythrocyte protein
 very abundant (340 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
Endocytosis of Host Cytoplasm
cytostome
food vacuole
pinocytosis
(rings)
The Food Vacuole
A Specialized Lysosome
ATP
hemoglobin
digestion
Endocytic
Pathway
H+
(pH 5-5.4)
Food Vacuole Proteases
• plasmepsins I - IV (acid)
• falcipains I - III (thiol)
• falcilysin (metallo)
Absent:
• other acid hydrolases
except acid phosphatase
ADP
parasite
cytoplasm
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
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
-F33/L34

• the large globin fragments
dissociate
– heme is released
• globin fragments are
susceptible to further
proteolysis
Hemoglobin Digestion is an
Ordered Process
hemoglobin
plasmepsin
large globin
+ heme
fragments
amino acids
aminopeptidase
di-peptides
falcipain
plasmepsin
dipeptidyl
aminopeptidase
medium fragments
small peptides
(~20 amino acids) falcilysin (6-8 amino acids)
aminopeptidase
amino acids
Membrane Transport
• Channel proteins (hydrophilic pores)
• Carrier proteins
– substrate specific
– most require energy
– ATPase or gradients
• 6 amino acid transporters identified in
Plasmodium genome (location?)
• PfMDR-1 and PfCRT located on food
vacuole membrane
PfMDR-1
• Member of ABC (ATP-binding cassette)
transporter super family
• Associated with drug resistance (MDR =
multi-drug resistance)
• Capable of peptide transport
– complements yeast ste6 gene (transporter
of yeast peptide mating factor)
• However, imports solutes (including
drugs) into food vacuole
PfCRT
• Member of DMT (drug/metabolite
transporter) super family
• Associated with chloroquine resistance
(CRT = chloroquine resistance
transporter)
• Exports chloroquine and other drugs
from the food vacuole
• Peptides can block drug export
The Food Vacuole
A Specialized Lysosome
ATP
hemoglobin
H+
proteins
plasmepsin
globin
heme +
fragments
ADP
 transporter s associated
with food vacuole
 amino-peptidase activities
in parasite cytoplasm
?
amino
acids
falcipain
amino
acids
plasmepsin
aminopeptidase
falcilysin
ATP
small fragments
(6-8 amino acids)
PfCRT?
ADP
Free Heme is Toxic
• heme destabilizes and lyses
membranes
• hydrolases released into
parasite cytoplasm
• parasite dies
Possible Detoxification Mechanisms
• heme  hemozoin (malaria pigment)
• H2O2 mediated degradation
• GSH mediated degradation
• heme oxygenase (P.b. and P.k. only)
Hemozoin = b-Hematin
heme
b-hematin
b-hematin forms insoluble crystals
'biocrystallization' or 'biomineralization'
Pigment Formation
• biocrystallization mechanism unknown
• b-hematin can form spontaneously (harsh
conditions)
• lipid bodies can promote the process
– derived from PVM
• potential heme detoxification protein
recently described
– unique to Plasmodium species
– binds 2-3 heme groups with high affinity (80 nM)
– exported to host cytoplasm and taken up into
food vacuole
• heme biocrystallization inhibited by
chloroquine and other anti-malarials
The Food Vacuole
A Specialized Lysosome
ATP
hemoglobin
O2
Fe3+
-O  O
2
2
Fe2+
plasmepsin
globin
heme +
fragments
falcipain
?
 iron oxidized after
release from Hb
 oxidation promotes
formation of ROS
 oxidative stress
H+
plasmepsin
hemozoin
falcilysin
ADP
amino
acids
aminopeptidase
ATP
small fragments
(6-8 amino acids)
ADP
The Food Vacuole
A Specialized Lysosome
ATP
hemoglobin
O2
Fe3+
-O  O
2
2
Fe2+
superoxide
dismutase?
H2O2
H+
plasmepsin
ADP
globin
heme +
fragments
?
amino
acids
falcipain
hemozoin
aminopeptidase
plasmepsin
falcilysin
ATP
catalase?
H2O + O2
amino
acids
small fragments
(6-8 amino acids)
?
ADP