GlycolysisGlycolysis

GluconeogenesisGluconeogenesis

Glycolysis - OverviewOne of best characterized pathways

Characterized in the first half of 20th century

Glucose --> 2 pyruvates + energy

Strategy

add phosphoryl groups to glucose

convert phosphorylated intermediates into compounds with high phosphate group-transfer potentials

couple the subsequent hydrolysis of reactive substances to ATP synthesis

Glucose + 2NAD+ + 2 ADP + 2Pi -->

2NADH + 2 pyruvates + 2ATP + 2H2O + 4H+

Overview of Glycolysis

The Embden-Meyerhof (Warburg) Pathway

• Essentially all cells carry out glycolysis

• Ten reactions - similar in most cells - but rates differ

• Two phases:

– First phase converts glucose to two G-3-P

– Second phase produces two pyruvates

• Products are pyruvate, ATP and NADH

• NADH must be recycled

• Three possible fates for pyruvate

Glycolysis

Glycolysis

Fate of pyruvate

Decarboxylation to acetaldehyde

Reduction to ethanol

Reduction to lactate

Mitochondrial oxidation

1 NADH --> ~3 ATP

Enzymes of glycolysisCatalyzed reactions

and properties

Enzymes of glycolysisCatalyzed reactions

and properties

Glucose

Glucose-6-phosphate

Fructose-6-phosphate

Fructose-1,6-biphosphate

Glyceraldehyde-3-phosphate

Hexokinase, glucokinase

Phosphoglucoisomerase

Phosphofructokinase

Aldolase

Triose phosphate isomerase

Dihydroxyacetone phosphate

First Phase of GlycolysisThe first reaction - phosphorylation of glucose

• Hexokinase or glucokinase • This is a priming reaction - ATP is consumed here

in order to get more later • ATP makes the phosphorylation of glucose

spontaneous

Hexokinase 1st step in glycolysis; G large, negative

• Hexokinase (and glucokinase) act to phosphorylate glucose and keep it in the cell

• Km for glucose is 0.1 mM; cell has 4 mM glucose

• So hexokinase is normally active!

• Glucokinase (Kmglucose = 10 mM) only turns on when

cell is rich in glucose• Hexokinase is regulated - allosterically inhibited by

(product) glucose-6-P -

Hexokinase

• First step in glycolysis• Large negative deltaG • Hexokinase is regulated - allosterically inhibited by

(product) glucose-6-P• Corresponding reverse reaction (Gluconeogenesis) is

catalyzed by a different enzyme (glucose-6-phosphatase)

• Is it the committed step in glycolysis ?

Glucose

Fructose-6-P

Glucose-6-P

Glyceraldehyde-3-P

Pyruvate

ATP

Glycogen Ribose-5-P + NADPH

Nucleic acidsynthesis

Reducingpower

Glucose-6-P dehydrogenase

Rx 2: Phosphoglucoisomerase

Glucose-6-P to Fructose-6-P

Rx 3: PhosphofructokinasePFK is the committed step in glycolysis!

• The second priming reaction of glycolysis • Committed step and large, neg delta G - means PFK is

highly regulated • ATP inhibits, AMP reverses inhibition • Citrate is also an allosteric inhibitor • Fructose-2,6-bisphosphate is allosteric activator • PFK increases activity when energy status is low • PFK decreases activity when energy status is high

Glycolysis - Second Phase

Metabolic energy produces 4 ATP

• Net ATP yield for glycolysis is two ATP

• Second phase involves two very high energy phosphate intermediates

• .

– 1,3 BPG

– Phosphoenolpyruvate

Glyceraldehyde-3-phosphate

1,3-biphosphoglycerate

3-phosphoglycerate

2-phosphoglycerate

phosphoenolpyruvate

pyruvate

Glyceraldehyde-3-phosphate dehydrogenase

Phosphoglycerate kinase

Phosphoglycerate mutase

Enolase

Pyruvate kinase

Rx 10: Pyruvate Kinase

PEP to Pyruvate makes ATP

• These two ATP (from one glucose) can be viewed as the "payoff" of glycolysis

• Large, negative G - regulation!

• Allosterically activated by AMP, F-1,6-bisP

• Allosterically inhibited by ATP and acetyl-CoA

The Fate of NADH and PyruvateAerobic or anaerobic??

• NADH is energy - two possible fates: – If O2 is available, NADH is re-oxidized in the

electron transport pathway, making ATP in oxidative phosphorylation

– In anaerobic conditions, NADH is re-oxidized by lactate dehydrogenase (LDH), providing additional NAD+ for more glycolysis

The Fate of NADH and PyAerobic or anaerobic??

• Pyruvate is also energy - two possible fates: – aerobic: citric acid cycle

– anaerobic: LDH makes lactate

The elegant evidence of regulation!

• Standard state G values are scattered: + and -

G in cells is revealing:

• Most values near zero

• 3 of 10 reactions have large, negative G

• Large negative G reactions are sites of regulation!

Energetics of Glycolysis

Gluconeogenesis

Synthesis of "new glucose" from common metabolites

• Humans consume 160 g of glucose per day

• 75% of that is in the brain

• Body fluids contain only 20 g of glucose

• Glycogen stores yield 180-200 g of glucose

• So the body must be able to make its own glucose

Comparison of glycolysis and gluconeogenesis pathways

Substrates for Gluconeogenesis

Pyruvate, lactate, glycerol, amino acids and all TCA intermediates can be utilized

• Fatty acids cannot!

• Most fatty acids yield only acetyl-CoA

• Acetyl-CoA (through TCA cycle) cannot provide for net synthesis of sugars

Gluconeogenesis I

• Occurs mainly in liver and kidneys

• Not the mere reversal of glycolysis for 2 reasons:– Energetics must change to make

gluconeogenesis favorable (delta G of glycolysis = -74 kJ/mol

– Reciprocal regulation must turn one on and the other off - this requires something new!

Energetics of Glycolysis

The elegant evidence of regulation!

G in cells is revealing:

• Most values near zero

• 3 of 10 reactions have large, negative G

• Large negative G reactions are sites of regulation!

• Reactions 1, 3 and 10 should be different to go into opposite direction

Gluconeogenesis II Something Borrowed, Something New

• Seven steps of glycolysis are retained:– Steps 2 and 4-9

• Three steps are replaced:– Steps 1, 3, and 10 (the regulated steps!)

• The new reactions provide for a spontaneous pathway (G negative in the direction of sugar synthesis), and they provide new mechanisms of regulation