FERMENTATION: Anaerobic
Glycolysis

CATABOLIC FATES OF PYRUVATE

HOMOLACTIC FERMENTATION
• Utilized by muscles when the demand for ATP is high and oxygen availability is low.
• ATP is rapidly regenerated compared to oxidative phosphorylation.
• The reaction is freely reversible

HOMOLACTIC FERMENTATION
• Net reaction:
Glucose + 2ADP+ 2Pi
2 lactate + 2ATP+ 2H
2
O+ 2H +
• Lactate formed can either exported from the cell or converted back to pyruvate
• The lactate formed in muscles is carried by the blood to the liver, where it is converted to glucose

CORI CYCLE

ALCOHOLIC FERMENTATION
• The NAD+ regenerated in this reaction will be utilized by GAPDH
• TPP is an important cofactor of Pyruvate decarboxylase

ALCOHOLIC FERMENTATION

GLYCOLYSIS AND CANCER
• Utilization of glucose and glycolysis proceed faster in cancer cells
• Because of hypoxia, cancer cells depend on anaerobic glycolysis for ATP production
• Tumor cells also have smaller amount of mitochondria
• Some tumor cells overproduce several glycolytic enzymes due to the presence of HIF-1
• HIF-1 acts at the level of mRNA synthesis to stimulate the production of at least 8 glycolytic enzymes

Entry of other sugars: Lactose

Entry of other sugars: Fructose

SYNTHESIS OF ACETYL Co-A
Pyruvate dehydrogenase complex is composed of 3 enzymes and requires 5 coenzymes

PYRUVATE DEHYDROGENASE
COMPLEX
• E1: pyruvate dehydrogenase (30 heterodimers)
• E2: dihydrolipoamide transacetylase (20 trimers)
• E3: dihydrolipoamide dehydrogenase (12 dimers)
• ~10,000 kD

PYRUVATE DEHYDROGENASE
COMPLEX

Thiamine = Vitamin B1
(ribo)Flavin = Vitamin B2
Niacin = Vitamin B3
Pantothenic Acid = Vitamin B5

REGULATION OF THE COMPLEX
• The eukaryotic complex contains two regulatory enzymes: a kinase that phosphorylates three serine residues in E1 and the phosphatase that removes those phosphates
• The activity of the complex is controlled by allosteric inhibition and covalent modification that is in turn controlled by the energy state of the cell.
• ATP is an allosteric inhibitor of the complex; AMP is an activator
• E2 is inhibited by acetyl-CoA and activated by CoA-SH
• E3 is inhibited by NADH and activated by NAD +

REGULATION OF THE COMPLEX
• Regulation also occurs by covalent modification of E1 (de/phosphorylation)
• NADH and acetyl-CoA activate the pyruvate dehydrogenase kinase which phosphorylates the 3 specific serine residues in E1 rendering it inactive
• Pyruvate dehydrogenase phosphatase removes the phosphate groups. This enzyme is activated by Ca 2+ and Mg 2+

KREBS CYCLE
• Aka tricarboxylic acid cycle and citric acid cycle
• central oxidative pathway
• Composed of 8 reactions that oxidizes acetyl
CoA to 2 molecules of CO2
• Occurs in the mitochondrial matrix

Citrate Synthase Reaction (First)
O
O O
O
O
H
2
O CoASH
O O
O
SCoA + acetyl CoA citrate synthase O
H O
O
O
O oxaloacetate citrate
• Claisen condensation
• OAA must bind first before Acetyl-CoA (sequential mechanism)
• -32.2kJ

Aconitase Reaction
O O
H O
O
O
O O citrate aconitase
H O
O
O
O
O
O
O isocitrate
• Forms isocitrate
• Goes through alkene intermediate (cis-aconitate)
– elimination then addition
• 13.3kJ

Isocitrate Dehydrogenase
H O
O
O
O
O
O
O isocitrate
NAD NADH CO
2 isocitrate dehydrogenase
O O
O
O
O alpha ketoglutarate
• All dehydrogenase reactions make NADH or FADH
2
• Oxidative decarboxylation
• -20.9kJ
• Energy from increased entropy in gas formation

α-ketoglutarate dehydrogenase
O O
O CoASH
CO
2 NAD
NADH
O
SCoA
O O alpha ketoglutarate alpha ketoglutarate dehydrogenase
• Same as pyruvate dehydrogenase reaction
• Formation of thioester
– endergonic
– driven by loss of CO
• increases entropy
2
• exergonic
• -33.5kJ
O O succinyl CoA

Succinyl CoA synthetase
O
SCoA
GDP
GTP CoASH
O O succinyl CoA succinyl CoA synthetase
• Hydrolysis of thioester
– Releases CoASH
– Exergonic
• Coupled to synthesis of GTP
– Endergonic
– GTP very similar to ATP and interconverted later
• -2.9kJ
O
O
O O succinate

Succinate dehydrogenase
O
O
FAD
FADH
2
O
O
H H succinyl CoA dehydrogenase
O O succinate
• Dehydrogenation
• Uses FAD
– NAD used to oxidize oxygen-containing groups
• Aldehydes
• alcohols
– FAD used to oxidize C-C bonds
– 0kJ
O O fumarate

Fumarase
H
2
O
H
O
O
O
H fumarate
O fumarase
• Addition of water to a double bond
• -3.8kJ
O
O
OH
O O malate

Malate Dehydrogenase
O
O
OH
O O malate
NAD NADH malate dehydrogenase
O
O
O
O
O oxaloacetate
• Oxidation of secondary alcohol to ketone
• Makes NADH
• Regenerates oxaloacetate for another round
• 29.7 kJ

REGULATION OF KREBS CYCLE
• 3 rate determining enzymes: citrate synthase, isocitrate dehydrogenase and alpha-ketoglutarate dehydrogenase
• 3 mechanisms used by the enzymes:
– Substrate availability (acetyl CoA and oxaloacetate)
– Product inhibition (NADH)
– Competitive feedback inhibition by intermediates (citrate and succinyl CoA
• ADP is an effector of isocitrate dehydrogenase
• Ca2+ activates pyruvate dehydrogenase phosphatase, isocitrate dehydrogenase and alpha-ketoglutarate dehydrogenase

Counting ATP’s: one molecule of glucose
ATP
Glycolysis 2
PDC (X2) 0
TCA (X2) 2
NADH FADH2
2
2
6
0
0
2
TOTAL 4
After
OxPhos
4
10 2
30 ATPs 4 ATPs