BIOS 301 - Fermentation & Pentose Phosphate Pathway

2-deoxyglucose

• Glycolysis occurs at elevated rates in tumor cells

• 2-deoxyglucose can be administered to cancer patients - enters glycolysis pathway but cannot proceed past second step

  • Limits energy production in cancer cells

  • But not a good drug - has to be given in toxically high doses to compete with glucose in glycolysis pathway

Pet scanning

• Administer 18F-fluorodeoxyglucose to patient

• Can be converted to FDG-6P but not metabolized further

  • Builds up quickly cells in with high metabolic activity (e.g. tumor cells)

• FDG-6P can be detected in PET scans to show location of cancer metastases

Fates of pyruvate (in animals)

• Heart muscle and brain: pyruvate is always subject to the TCA cycle and complete oxidation to ensure a constant supply of ATP reaches vital tissues

• Skeletal muscle: when this tissue goes anaerobic, lactic acid fermentation occurs

• Liver: can conduct lactic acid fermentation in reverse (gluconeogenesis) to ensure energy derived from glucose can be maximized

Purpose of fermentation

• Generates ATP under anaerobic conditions

• Recycles NADH → NAD+ for further glycolysis (i.e. energy production)

Lactic acid fermentation

• 2 pyruvate + 2 NADH → 2 lactate + 2 NAD+

• Catalyzed by lactate dehydrogenase

• Occurs in an oxidation-reduction reaction

Reduction potential

• High reduction potential corresponds to high affinity for electrons

  • Electrons are transferred from lower to higher E

• Delta E = Eacceptor -Edonor

  • Delta E must be positive to be thermodynamically favorable (negative delta G)

• To find delta E use half reactions of pyruvate (acceptor) and NADH (donor)

Isozymes of lactate dehydrogenase

• Muscle LDH has higher affinity for pyruvate than heart LDH

  • Allows for shorts bursts of energy by anaerobic glycolysis in muscle

  • Forces aerobic, complete oxidation of glucose in the heart

    • Heart LDH is also subject to excess substrate inhibition

  • There are two genes for LDH monomers (H and M) which combine randomly to form tetramer enzyme molecules

    • Blood plasma tests can reveal identity of injured/necrotic tissue based on LDH subunit representation

Ethanol fermentation (step 1)

• Pyruvate → Acetaldehyde + CO2

• Enzyme: pyruvate decarboxylase

• Cofactors: Mg2+, TPP

Ethanol fermentation (step 2)

• Acetaldehyde + NADH → Ethanol + NAD+

• Enzyme: alcohol dehydrogenase

TPP

• Thiamine pyrophosphate

• Contains thioazolium ring: allows for electron delocalization → generates carbanion that attacks substrate → facilitates C-C bond cleavage and decarboxylation

Pentose Phosphate Pathway (overall result)

• Substrate is glucose-6-phosphate

• Main products are NADPH and ribose-5-phosphate

NADPH purpose

• Reductive biosynthesis of fatty acids and steroids

• Repair of oxidative damage

• Reduction of glutathione

Ribose-5-phosphate purpose

Used in synthesis of DNA/RNA or certain coenzymes

Pentose Phosphate Pathway (substrates, products, enzymes)

Oxidative vs. Nonoxidative phase of PPP

• Oxidative phase yields NADPH and ribulose-5-phosphate

• Nonoxidative phase: if ribulose-5-phosphate is already abundant, can be converted to xylulose-5-phosphate and back to G-6-P

  • Generates NADPH while conserving glucose

Transketolases

• Assist in nonoxidative phase of PPP

• Transfer 2C units from ketose donor → aldose acceptor

• Reaction intermediate stabilized by TPP

Transaldolases

• Assist in nonoxidative phase of PPP

• Transfer 3C units from ketose donor → aldose acceptor

• Reaction intermediate is a protonated Schiff’s base

Partitioning into glycolysis vs. PPP

If NADPH is sufficiently high, PPP is inhibited, and glycolysis proceeds instead

G-6-P Dehydrogenase Deficiency

• G-6-P Dehydrogenase is responsible for recycling NADP+ into NADPH

• NADPH is required by glutathione reductase to convert oxidized glutathione → reduced form

• Glutathione is necessary to neutralize reactive oxygen species

PPP in tumor cells

G-6-P Dehydrogenase, 6-phosphogluconolactonase, and 6-phosphogluconate dehydrogenase are all upregulated in tumor cells