Glycolysis • What is glycolysis? • sequence of reactions that converts one molecule of glucose to two molecules of pyruvate with the formation of two ATP molecules • anaerobic
Glycolysis • Why is glucose such a commonly used fuel? • tends to exist in ring form, very stable, doesn’t generally glycosylate proteins • formed from formaldehyde under prebiotic conditions
Glycolysis • What are the possible fates of glucose?
Glycolysis • What’s the difference between a facultative anaerobe and an obligate anaerobe? • Can you give an example of habitat-dependent anaerobiosis? • What about activity-dependent anaerobiosis?
Glycolysis • All the intermediates in glycolysis have either 3 or 6 carbon atoms • All of the reactions fall into one of 5 categories • phosphoryl transfer • phosphoryl shift • isomerization • dehydration • aldol cleavage
Glycolysis • Entire reaction sequence may be divided into three stages • glucose is trapped and destabilized • six carbon molecule is split into two three carbon molecules • ATP is generated
Glycolysis – Stage 1 • glucose converted to glucose-6-PO4 • ATP is needed • catalyzed by hexokinase or glucokinase • induced fit • G01= -4.0 kcal/mole
Glycolysis – Stage 1 • phosphoglucoisomerase • aldose is converted to ketose • G01=+0.4 kcal/mole
Glycolysis – Stage 1 • rate limiting enzyme – allosteric • inhibited by high ATP, citric acid, long-chain fatty acids • stimulated by ADP or AMP • G01= - 3.4 kcal/mole
Glycolysis – Stage 2 • six carbon molecule split into 2- 3 carbon molecules • aldose and ketose • G01=+ 5.73 kcal/mole
Glycolysis – Stage 3 • At equilibrium most mixture exists as dihydroxyacetone phosphate • G01=+ 1.83 kcal/mole
Glycolysis – Stage 3 • redox reaction • energy from redox used to form acyl phosphate • G01= +1.5 kcal/mole
Glycolysis – Stage 3 • Consists of two coupled processes
Glycolysis – Stage 3 • formation of ATP – substrate level phosphorylation
Glycolysis – Stage 3 • phosphoryl shift – uses 2,3 bisphosphoglycerate G01= +1.1 kcal/mole • dehydration G01 = +.44 kcal/mole • phosphoryl transfer G01 = -7.5 kcal/mole
Alcoholic Fermentation • Which organisms carry out this process? • yeast • other microorganisms • PDC requires thiamine pyrophosphate as coenzyme • NAD+ is regenerated
Lactic Acid Fermentation • Occurs in muscle cells, microorganisms • Regenerates NAD+
NAD+ and Dehydrogenases • Various dehydrogenases have a similar binding domain for NAD+ showing their common origin • Rossman fold
Glycolysis • How can fructose be used for energy?
Glycolysis • To use galactose it must be converted to glucose-6-PO4
Glycolysis • What causes lactose intolerance?
Glycolysis • What is galactosemia? • inability to metabolize galactose • missing galactose 1-phosphate uridyl transferase • liver disease • development of cataracts • CNS malfunction
Control of Glycolysis • Of what value is glycolysis for cells? • provides energy in form of ATP • provides building blocks for synthetic reactions • Where are most control points found? • enzymes that catalyze irreversible reactions • hexokinase • phosphofructokinase • pyruvate kinase
Phosphofructokinase • Most important control point in mammalian glycolytic pathway • allosteric enzyme • activated by AMP and fructose 2,6 bisphosphate • inhibited by high levels of ATP, citrate, fatty acids
Hexokinase • Hexokinase is inhibited by its product glucose-6-PO4 • glucose remains in blood • Glucokinase, an isozyme of hexokinase is not inhibited by glucose-6-PO4 • found in liver • has lower affinity for glucose
Pyruvate Kinase • Pyruvate kinase exists as isozymes • L form – predominates in liver • M form – mostly in muscle and brain • PK is an allosteric enzyme • activated by fructose 1,6 bisphosphate • inhibited by ATP, alanine • L form of PK influenced by covalent modification • inhibited by phosphorylation
Glucose Transport • What is the role of glucose transporters in animal cells? • facilitate movement of glucose across cell membrane • What kind of molecule is a transporter and where is it located? • small protein embedded in plasma membrane
Glucose Transport • mammalian glucose transporter
Glycolysis and Cancer • Why are rapidly growing tumor cells dependent upon glycolysis? • insufficient oxygen supply • What is the function of HIF-1? • hypoxia-inducible transcription factor stimulates synthesis of many glycolytic enzymes and GLUT-1 and 3 • also stimulates vascular endothelial growth factor
Gluconeogenesis • What is gluconeogenesis? • synthesis of glucose from non-carbohydrate precursors • Why is this an important pathway? • What are some of the major precursors? • lactate, amino acids, glycerol • Where does this process occur? • liver, kidney
Gluconeogenesis • If gluconeogenesis involves the conversion of pyruvate to glucose why is it not simply the reverse of glycolysis? • glycolysis contains several irreversible reactions • Which reactions in glycolysis are irreversible? • phosphoenolpyruvate to pyruvate • fructose 6-phosphate to fructose 1,6-bisphosphate • glucose to glucose 6-phosphate
Gluconeogenesis • What is the first reaction?
Gluconeogenesis • Why is pyruvate carboxylase of special interest? • structural properties • contains ATP-grasp domain at N-terminal end • contains biotin-binding domain at C-terminal end
Gluconeogenesis • What is the role of biotin in this reaction? • prosthetic group lined to -amino group of lysine residue • carrier of activated carbon dioxide
Gluconeogenesis • Pyruvate carboxylase is an allosteric enzyme • activated by acetyl CoA • needed to form carboxybiotin
Gluconeogenesis • Carboxylation of pyruvate occurs in the mitocondria but next step in reaction sequence occurs in cytosol
Gluconeogenesis Decarboxylation of oxaloacetate is coupled with phosphorylation by GTP enzyme is phosphoenolpyruvate carboxykinase
Gluconeogenesis • Which other steps in glycolysis are irreversible? • conversion of fructose 1,6-bisphosphate to fructose 6-phosphate • conversion of glucose 6-phosphate to glucose
Gluconeogenesis • G° = -16.7 kJ mol-1 • fructose-1,6-bisphosphatase is an allosteric enzyme, inhibited by AMP and activated by ATP
Gluconeogenesis • Enzyme that catalyzes last reaction not found in all tissues • liver and kidney cortex
Gluconeogenesis • Is gluconeogenesis an energetically favorable reaction in the cell? • What drives this reaction? • Are glycolysis and gluconeogenesis active at the same time?