glycolic processes Flashcards
Which proteins are usually used to transport glucose into cells?
- GLUT proteins
When are GLUT 1 and GLUT 2 proteins used for transport and how do they compare?
- often GLUT 1
- not modulated by insulin
- has high affinity for glucose (low Km) but low capacity
- GLUT 2 used by hepatocytes
- low affinity for glucose (high Km), but high capacity
- regulated by insulin – forms part of glucose-sensing system w/ glucokinase
- phosphorylated by hexokinase to maintain cellular glucose absorption (glucose to G6P)
When is GLUT 4 used and why?
- in muscle and adipose tissue
- insulin-regulated with a relatively high glucose affinity (low Km)
- a facilitative transporter
What is normal BGC when fasted?
4-7mmol/L
Describe the metabolism of glucose by glycolysis.
- glucose breakdown (6 carbons) to 2x pyruvate (3 carbons)
- both use and production of ATP (energy)
- also production of reducing equivalents for energy-production in oxidative phosphorylation
- located in cytosol
Describe reactions 1-5 of glycolysis with the:
- substrate → product (enzyme)
- the point of the reaction
1) glucose → glucose-6-phosphate (hexokinase)
- traps glucose in cells and destabilises structure to facilitate later reactions
2) glucose-6-phosphate → fructose-6-phosphate (phosphoglucose isomerase)
- converts 6C ring into a 5C ring in preparation for triose formation
3) fructose-6-phosphate → fructose-1,6-bisphosphate (phosphofructokinase)
- further destabilisation of structure, preparation for triose formation
4) fructose-1,6-bisphosphate → dihdroxyacetone/glyceraldehyde-3-phosphate
(aldolase)
- splits 5C ring into 2x triose sugars
5) dihdroxyacetone → glyceraldehyde-3-phosphate (triosephosphate isomerase)
- isomerisation reaction as only G-3-P can proceed for future reactions
Describe reactions 6-10 of glycolysis with the:
- substrate → product (enzyme)
- the point of the reaction
6) glyceraldehyde-3-phosphate → 1,3-bisphosphoglycerate (glyceraldehyde phosphate dehydrogenase)
- to provide 2x phosphate groups for ATP synthesis in subsequent reactions
7) 1,3-bisphosphoglycerate → 3-phosphoglycerate (phosphoglycerate kinase)
- substrate-level phosphorylation to produce ATP
8) 3-phosphoglycerate → 2-phosphoglycerate (phosphoglycerate mutase)
- isomerism to promote the formation of more unstable phosphoenolpyruvate
9) 2-phosphoglycerate → phosphoenolpyruvate (enolase)
- formation of unstable product for next rxn
10) phosphoenolpyruvate → pyruvate (pyruvate kinase)
- substrate-level phosphorylation to produce ATP
How do glycolysis phase 1 (rxns 1-5) and glycolysis phase 2 (rxns 6-10) compare?
- phase 1 – uses 2x ATP
- phase 2 – uses no ATP
State the total energy balance of glycolysis in terms of phase 1 and phase 2.
- 2x triose sugars (G-3-P) produced in phase 1 • phase one - ATP: -2 - NADPH: 0 • phase two - ATP: +4 - NADPH: +2
TOTAL = 2x ATP and 2xNADPH
How does glycolysis occur with galactose and fructose as a pose to glucose?
- galactose converted by multi-step pathway to glucose-6-phosphate and then enters glycolysis
- fructose phosphorylated by hexokinase (muscle + adipose tissue) to fructose-6-phosphate and then enters glycolysis
Describe regulation and control of glycolysis.
- most bodily reactions reversible but 3 steps of glycolysis irreversible because of energy input by ATP
- reversible reactions instead regulated via concentrations of substrates and products
- enzymes regulated by allosteric regulation, hormonal signalling action (short-term) and induction/repression of enzyme synthesis (long-term)
Compare hexokinase and glucokinase activity.
• hexokinase
- universal (most cells)
- inhibited by glucose-6-phosphate (G6P)
- unaffected by insulin
- low Km (0.01 mM glucose)
- will phosphorylate other sugars
• glucokinase
- liver & kidney
- no effects of G-6-P
- regulated by insulin + glucagon
- high Km (12mM glucose)
- specific for glucose
Describe glycolysis regulation by phosphofructokinase (PFK)
• inhibited by: - ATP - citrate - glucagon • stimulated by: - AMP - insulin
Describe glycolysis regulation by pyruvate kinase (PFK)
• inhibited by: - ATP - acetyl CoA - glucagon • stimulated by: - fruct-1,6-bisphos - insulin
Compare where the NAD⁺ needed for pyruvate reactions comes from in aerobic and anaerobic conditions.
• aerobic conditions:
- NAD⁺ regenerated in mitochondria (oxidative phosphorylation)
- NAD⁺/NADH cannot cross mitochondrial membrane
- glycerin-phosphate shuttle pathway
- malate-aspartate shuttle pathway
• anaerobic conditions:
- lack of oxidation of NADH to NAD⁺ in mitochondria
- reduction of pyruvate to lactate used to produce NAD⁺
- oxidation of malate to oxaloacetate (intermediate in Krebs cycle)
Compare the glycerin-phosphate and malate-aspartate shuttle.
- dihydroxygacetone-phosphate (NADH to NAD⁺) → glycerin-3-phosphate → (FAD → FADH₂) →
dihydroxygacetone-phosphate - oxaloacetate → aspartate → oxaloacetate → malate (NADH to NAD⁺) → malate (NADH to NAD⁺) → oxaloacetate
(see notes for diagrams)
Describe the fates of pyruvate.
- (aerobic) pyruvate enters Krebs cycle, producing NADH, FADH₂ and GTP
- (anaerobic) pyruvate can either be converted to lactate or fermented to ethanol (in microorganisms)
- both processes use up NADH, converting it to NAD⁺
- key in allowing regeneration of NAD⁺ to continue glycolysis
What is gluconeogenesis and where does it occur?
- formation of glucose from non-carbohydrate sources
- mainly in liver (not all cells do this)
- 3 irreversible steps - must be worked around
- other reactions are close to equilibrium - concentrations of substrates and products determines direction of flow
(see notes for diagrams)
State some gluconeogenic substrates.
- AAs (not leucine or lysine)
- lactate
- pyruvate
- glycerol only (from stored fats)
- oxaloacetate
NB: 2 reactions that produce oxaloacetate from pyruvate (2 molecules of p = 1 molecule of glucose)
Describe the reaction in gluconeogenesis which starts with triglyceride.
Triglyceride → glycerol → glyceraldehyde-3-phosphate → fructose 1,6-bisphosphate
triglyceride ⬇ free FAs ⬇ acetyl CoA
(gylcerol can enter fluconeogenesis, animals cannot produce pyruvate from actyel-CoA and glucose cannot be synthesised from FAs)
Describe the conversion of pyruvate to phosphoeonolpyruvate (PEP).
- pyruvate (pyruvate decarboxylase) → oxaloacetate (phosphoeonolpyruvate carboxykinase) → PEP
- requirement for ATP
- GTP hydrolysis for substrate-level phosphorylation
- pyruvate carboxylase is mitochondrial enzyme so must transport oxaloacetate out of mitochondria
- mechanism depends on how cell regenerates NADH for use in G-3-P dehydrogenase catalysed reaction
Compare conversion of pyruvate to PEP under normal conditions and under stress (during exercise).
• normal conditions:
- pyruvate transported into mitochondria
- pyruvate (pyruvate carboxylase) → oxaloacetate (reduced) → malate
- malate exported to cytosol (using NADH)
- malate (oxidised) → oxaloacetate (cytosolic malate dehydrogenase) + NADH (regenerated)
- oxaloacetate → PEP
• unders stress/exercise:
- lactate → pyruvate + NADH (regenerated for GAPDH reaction use)
- in mitochondria: pyruvate → PEP (exported to cytosol)
Describe the stages of gluconeogenesis.
pyruvate → oxaloacetate → PEP → 2-phosphoglycerate → 3-phosphoglycerate → 1,3-bisphosphoglycerate (up to here requires 2xATP per reaction) → glyceraldehyde 3-phosphate (requires 2x ADH)→ dihydroxyacetone phosphate → fructose 1,6-phosphate → fructose 6 phosphate → glucose-6-phosphate → glucose
(see notes for detailed diagram)
Describe allosteric regulation of gluconeogenesis.
• energy status indictaor metabolites reciprocally-regulate enzymes of glycolysis + gluconeogenesis
- i.e. AMP stimulates PFK (glycolytic) while also inhibiting fructobisphosphatase-1 action (gluconeogenesis)
• also specific metabolites activating/inhibiting particular enzymes non-reciprocally
- i.e. ATP action on PFK
• fructose-2,6-bisphosphate also an important specific allosteric regulator
Describe fructose-2,6-bisphosphate.
- synthesised from fructose-6-phosphate
- allosteric binding to enzymes results in increased/decreased affinity for substrates
- provides mechanism modification of glycolysis/gluconeogenesis via insulin + glucagon signalling which results in dephosphorylation/phosphorylation of dual-function enzyme called PFK2
- high BGC results in removal of Ser-32 residue allowing hormonal regulation
- PFK2 catalyses both synthesis or degradation: fructose-2,6-bisphosphate ⇌fructose-6-phosphate
What is the importance of metabolite transport between tissues?
- glycolysis found in almost all cell types in body
- gluconeogenesis occurs primarily in liver
- thus, organs and tissues which produce lactate (e.g. hypoxic tissue) cannot convert it back to glucose
- transporting these metabolic products like to liver, can be recycled back to glucose and stored in the body by Cori cycle
What is the energy cost of the Cori cycle?
- net 2x ATP produced in anaerobic glycolysis
- however, 4x ATP and 2x GTP used in gluconeogenesis = net 4x ATP (equivalents) used
What are the benefits of the Cori cycle?
- rapid post-exercise replenishment of glycogen stores
- NAD⁺ regeneration for glycolysis + NADH regeneration for gluconeogenesis (in different tissues)
Describe the properties of glycogen.
- insoluble glucose polymer → carbohydrate storage
- monomers linked by α-1,4 bonds (straight chains) and α-1,6 bonds for branch points
- heavily-hydrated
- glycogenin core
- highly-branched
- allows improved solubility and sites for synthesis + degradation interactions so they can rapidly occur
Which two main tissues store glycogen and for what functions?
- liver (glucose-6-phosphatase)
- BGC maintenance
- released over long periods
- G-6-P → glucose - muscle
- energy provision
- released when instantaneously-required
Describe the 5 stages of glycogen catabolism, including enzymes.
- conversion: glucose → glucose-1-phosphate (glycogen phosphorylase)
- addition: UDP-Glucose + glycogen (via α-1,4 bond) (glycogen synthase)
- addition of glucose, via α-1,4 linkage, to non-reducing end of growing glycogen chains
- released UDP regenerated to UTP for use in glucose activation - reversal of above for catabolism (glycogen branching + debranching enzymes)
- conversion: glucose-1-phosphate → glucose-1-phosphate (phosphoglucomutase)
- ATP generates UTP: UDP → UTP + Pi
- UTP + G-1-P → UDP-glucose + PPi (UDP-glucose pyrophosphorylase)
- phosphorylation: ATP + glucose → G-1-P (traps glucose in cell)
- for every 1x mole of glucose = 2x moles ATP consumed in this step - activation of glucose: UDP + G-1-P → glucose (UDP-glucose pyrophosphorylase)
What are reducing and non-reducing ends of glucose?
- in solution, glucose ring structure is dynamic
- at C1, reducing end, there is formation of aldehyde group (oxidation possible) when open
- at C4, non-reducing end on other side of this structure
(see notes for diagrams)
Describe branching enzyme and its activity.
- or amylo-(1-4→1-6) transglycosylase
- adds branches to glycogen (via glucose units)
- after 10 glucose units added to glycogen, an α-1,6 branch point formed (branching enzyme)
- enzyme breaks one of the α-1,4 glycosidic bonds and transfers block of residues (about 7) to more interior site in glycogen
- these are then re-attached by an α-1,6 glycosidic bond
(see notes for diagram)
What is glycogenolysis requires for and which 3 enzymes are involved.
- catabolic process resulting in the formation of free glucose or glucose-6-phosphate
- 3 enzymes required:
1. glycogen phosphorylase
2. transferase enzyme
3. glycogen-debranching enzyme
Describe the process of glycogen breakdown.
- glycogen phosphorylase activated by: energy demand (AMP), muscle contraction (Ca2⁺ ions) and stress (adrenaline + glucagon)
- glycogen (non-reducing end) + pi (phosphate) G-1-P (breaks α-1,4 glycosidic bonds)
- processive enzyme - attaches to glycogen and removes glucose residues until >5 away from branchpoint
- remaining residues added to existing chain with α-1,4 glycosidic bond (transferase enzyme)
- residue at the α-1,6 branching point removed (glycogen-debranching enzyme)
- debranching enzyme has dual activity (transferase and debranching)
Describe the regulation of glycogen metabolism.
- G-1-P cannot directly enter glycolysis so phosphoglucomutase converts it to G-6-P first
- glucose-6-phosphate stays inside cell (ionised and saves energy) and can go straight into glycolysis
- glycogen phosphorylase degrades glycogen by breaking α-1,4 glycosidic bonds to release glucose units 1 at a time from non-reducing end of a glycogen chain (end with free 4’-OH group)
- glucose released as glucose-1-phosphate:
(glycogen) n + Pi ⇌ (glycogen)n-1 + glucose-1-phosphate - differences in muscle and liver glucagon acts on liver only - in liver cells glycogen phosphorylase is inactivated by high BGC
Describe glycogen synthesis.
- enzyme glycogen synthase inactivated by phosphorylation
- achieved via kinase cascade activation by hormones epinephrine and glucagon
- insulin signalling results in phosphatase activation
- this de-phosphorylates glycogen synthase and activates enzyme
Summarise hormonal control of glycogen synthesis.
- glycogen degradation or synthesis occurs depending on low or high BGC
- to prevent futile cycles and produce appropriate responses
(see notes for diagram summary)
Describe glycogen storage diseases which affect the liver only.
including their:
- deficient enzyme
- symptoms
• Von gierke:
- glucose-6-phosphatase
- increased glycogen stores, enlarged liver, kidney failure, hypoglycaemia
• Hers:
- liver glycogen phosphorylase
- increased glycogen stores, hypoglycaemia
• Type 0:
- glycogen synthase
- hypoglycaemia, ketosis, failure to thrive
• Type VIII/IX:
- phosphorylase kinase
- enlarged liver, lack of response to glucagon and epinephrine
Describe glycogen storage diseases which affect the liver and muscle, just muscle or all organs.
including their:
- deficient enzyme
- symptoms
• McArdle: (muscle only)
- liver glycogen phosphorylase
- moderate increase in muscle glycogen, exercise-induced cramps
• Cori: (liver + muscle)
- debranching enzyme (Cori cycle affected)
- enlarged liver, mild hypoglycemia
• Andersen: (liver + muscle)
- branching enzyme
- enlarged liver, failure to thrive (physically develop), not fatal
• Pompe: (all organs)
- lysosomal α-1,4-gucosidase
- herat failure in infantile form, muscle defects in juvenile form
