Lecture

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Lecture 7Pentose Phosphate Pathway

• Nisson Schechter PhD
• Department of Biochemistry and Cell Biology
• Department of Psychiatry
• HSC T10, Room 050/049
• Telephone [phone]
• FAX [phone]
• nisson.schechter_at_stonybrook.edu

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For ?G, the free energy of a reaction, standard
conditions were defined concentration of
reactants and products at 1M temperature at
25C acidity at pH 7.0 Under these standard
conditions, ?G0' is defined as the standard free
energy change.
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Reading Material
Any Biochemistry Textbook - Stryer, Lenninger
Web Sites
http//www.tcd.ie/Biochemistry/IUBMB-Nicholson/swf
/glycolysis.swf
http//www.northland.cc.mn.us/biology/Biology1111/
animations/glycolysis.html
http//www.biocarta.com/pathfiles/h_glycolysisPath
way.asp
http//www.accessexcellence.org/RC/VL/GG/out_Glyco
l.html
Powerpoint - on course web site
Credits
Nisson Schechter PhD Department of Biochemistry
and Cell Biology, Stonybrook, NY
Robert Roskoski, PhD Department of Biochemistry,
LSUHSC - NO
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Pentose Phosphate Pathway

• Marks Chap. 28/pp. 437- 444.
• Stryer Chap. 20/pp. 563 - 574.
• Lehninger Chap. 15/pp. 558 - 560.
• (Lehninger, not recommended for this topic)

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Pentose Phosphate Pathway

• Also known as
• Pentose shunt
• Hexose monophosphate shunt
• Phosphogluconate pathway
• It occurs in the cytosol.

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One fate of G6P is the pentose pathway.
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The pentose pathway is a shunt.

• The pathway begins with the glycolytic
  intermediate glucose 6-P.
• It reconnects with glycolysis because two of the
  end products of the pentose pathway are
  glyceraldehyde 3-P and fructose 6-P two
  intermediates further down in the glycolytic
  pathway.
• It is for this reason that the pentose pathway is
  often referred to as a shunt.

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Moderate glucose flux
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Large glucose flux
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Its a shunt
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What does the pentose phosphate pathway achieve?

• The pathway yields reducing potential in the form
  of NADPH to be used in anabolic reactions
  requiring electrons.
• The pathway yields ribose 5-phosphate.
• Nucleotide biosynthesis leading to
• DNA
• RNA
• Various cofactors (CoA, FAD, SAM, NAD/NADP).

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NADPH is a phosphorylated form of NADH.
In general, with some exceptions, NADH is used to
drive the phosphorylation of ADP to ATP. NADPH
is used where reducing potential is required for
synthetic reactions.
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The pentose pathway can be divided into two
phases.
Non-oxidative interconversion of sugars
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NADPH H is formed from two separate
reactions. The glucose 6-phosphate DH (G6PD)
reaction is the rate limiting step and is
essentially irreversible. There is a medical
story for this enzyme. Cells have a greater need
for NADPH than ribose 5-phosphate.
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Regulatory enzyme
5 carbon atoms
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Regulatory enzyme
The enzyme is highly specific for NADP the Km
for NAD is 1000 greater than for NADP.
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Dont panic, you need not know all the reactions
in detail stay tuned.
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The nonoxidative phase of the pentose pathway

• This entails extensive carbon atom rearrangement.

Transketolase requires the coenzyme thiamine
pyrophosphate (TPP), the transaldolase does not.
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• Transketolase (TPP) and transaldolase are the
  link back to glycolysis.
• Glyceraldehyde 3-phosphate
• Fructose 6-phosphate
• Net result
• 3C5 ? 2C6 C3

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Ingested ribose can enter the glycolytic pathway
through the pentose pathway.
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Regulation of the Pentose Pathway

• Glucose 6-phosphate DH is the regulatory enzyme.
• NADPH is a potent competitive inhibitor of the
  enzyme.
• Usually the ratio NADPH/NADP is high so the
  enzyme is inhibited.
• But, with increased demand for NADPH, the ratio
  decreases and enzyme activity is stimulated.

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• The reactions of the non-oxidative portion of the
  pentose pathway are readily reversible.
• The concentrations of the products and reactants
  can shift depending on the metabolic needs of a
  particular cell or tissue.

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Rapidly dividing cells require more ribose 5-
phosphate than NADPH.
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The need for NADPH and ribose 5-phosphate is
balanced.
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More NADPH is needed than ribose 5-phosphate
Fatty acid synthesis in adipose cells.
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The cell needs both NADPH and ATP
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Glutathione and NADPH

• What is glutathione?
• Why is it important?
• How is it related to NADPH?

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Glutathione is a tripeptide composed of
glutamate, cystein, glycine.
Reduced glutathione (GSH) maintains the normal
reduced state of the cell.
Reduced glutathione (GSH)
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Glutathione Functions -1

• It serves as a reductant.
• Conjugates to drugs making them water soluble.
• Involved in amino acid transport across cell
  membranes.
• Cofactor in some enzymatic reactions.
• rearrangement of protein disulfide bonds.

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Glutathione Functions -2

• The sulfhydryl of GSH is used to reduce peroxides
  (ROS) formed during oxygen transport.
• Reactive oxygen species (ROS) damage
  macromolecules (DNA, RNA, and protein) and
  ultimately lead to cell death.
• The resulting oxidized form of GSH is two
  molecules linked by a disulfide bridge (GSSG).

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The enzyme glutathione reductase uses NADPH as a
cofactor to reduce GSSG back to two moles of GSH.
Thus, the pentose pathway is linked to the
supply of adequate amounts of GSH.
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So, what happens if glucose 6-phosphate DH is
defective? Insufficient production of
NADPH. Which translates into insufficient
glutathione. Is this a medical problem? YES
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Glutathione and Erythrocytes -1

• GSH is extremely important particularly in the
  highly oxidizing environment of the red blood
  cell.
• Mature RBCs have no mitochondria and are totally
  dependent on NADPH from the pentose phosphate
  pathway to regenerate GSH from GSSG via
  glutathione reductase.
• In fact, as much as 10 of glucose consumption,
  by erythrocytes, is mediated by the pentose
  pathway.

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Glutathione and Erythrocytes -2

• The reduced form of glutathione serves as a
  sulfhydryl buffer.
• It maintains cysteine residues in hemoglobin and
  other proteins in a reduced state.
• GSH is essential for normal RBC structure and
  keeping hemoglobin in Fe state.

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Glutathione and Erythrocytes -3

• Reduced glutathione also detoxifies peroxides.
• 2GSH ROOH ? GSSG H2O ROH
• Cells with low levels of GSH are susceptible
  hemolysis.
• Individuals with reduced GSH are subject to
  hemolysis.
• This is often clinically seen as black urine
  under certain conditions.

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Conditions for hemolytic anemia related G6PD
deficiency.

• The ingestion of oxidative agents that generate
  peroxides or reactive oxygen species (ROS).
• Antimalarials - pamaquine
• purine glycoside from fava beans.
• Individules with G6PD deficiency can not produce
  sufficient GSH to cope with the ROS.
• Proteins become cross linked leading to Heinz
  body formation and cell lysis.

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Glucose 6-phosphate DH deficiency and
nonspherocytic hemolytic anemia.

• Over 300 genetic variants of the G6PD protein are
  known.
• Thus, there is a remarkable variation in the
  clinical spectrum.
• G6PD deficiency is an inheritable X-linked
  recessive disorder.
• Approximately 10-14 of the male African American
  population is affected.
• It is also seen in Caucasians from the
  Mediterranean Basin.

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• People with the disorder are not normally anemic
  and display no evidence of the disease until the
  red cells are exposed to an oxidant or
  stress.Drugs that can precipitate this reaction
• antimalarial agents
• sulfonamides (antibiotic)
• aspirin
• nonsteroidal antiinflammatory drugs (NSAIDs)
• nitrofurantoin
• quinidine
• quinine
• exposure to certain chemicals - mothballs

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FAVISM

• Individuals with G6PD deficiency must not eat
  Fava beans.
• Pythagoras
• Erythrocytes lysedark or black urine.
• Interesting
• The growth Plasmodium falciparum (malaria
  parasite) fails in G6PD deficient individuals.