Additional Pathways of Carb Metabolism Flashcards
What is gluconeogenesis?
The synthesis of glucose from non-carbohydrate precursors
Almost the reverse of glycolysis as it can by the pathway from pyruvate to glucose
Mainly occurs in the liver (only a little in the kidney)
What are the non-carbohydrate precursors that can be converted back into glucose?
Lactate, pyruvate, citric acid cycle intermediates and the carbon skeletons of most amino acids
What are the non-carbohydrate precursors initially converted into?
A common intermediate - oxaloacetate
(however, leucine and lysine cannot be converted to oxaloacetate, as their breakdown will produce acetyl-CoA)
Oxaloacetate is also a citric acid cycle intermediate
COO CH2 CO COO
Which stages of gluconeogensis are different in reverse to glycolysis?
Pyruvate -> phosphoenolpyruvate
Fructose-1,6-bisphosphate -> fructose-6-phosphate
Glucose-6-phosphate -> Glucose
What is step 1 of gluconeogensis?
Pyruvate is converted into phosphoenolpyruvate in two steps
Pyruvate -> oxaloacetate
Uses pyruvate carboxylase and ATP
Oxaloacetate -> phosphoenolpyruvate (PEP)
uses PEPCK and GTP
This is exothermic decarboxylation as CO2 is released
Step 1 gluconeogenesis - describe pyruvate carboxylase?
Tetrameric protein
Uses a biotin cofactor to facilitate catalysis
Biotin functions as a CO2 carrier
Biotin is a cofactor derived from vitamin B7 or H
Step 1 gluconeogenesis - what does pyruvate carboxylase do?
It has a two phase reaction:
Cleavage of ATP drives dehydration of bicarbonate to form high energy carboxyphosphate
Resulting CO2 reacts with biotin to yield an activated intermediate
Activated carboxyl group is transferred to pyruvate to form oxaloacetate
Two reaction phases occur on different sub-sites of the enzyme
The CO2 carrying biotin ring is transferred between the sites on its flexible arm
Give an overview of the enzyme positions in step 1 of gluconeogenesis?
Gluconeogenesis requires metabolite transfer between mitochondria and cytosol
Generation of oxaloacetate occurs in the mitochondria
The enzymes that convert PEP to glucose are cytosolic
Either the oxaloacetate or PEP must leave the mitochondria
PEP is transported by specific transporter systems
Oxaloacetate transport is more complex as it can’t cross the mitochondrion membrane
How is oxaloacetate transported out the mitochondrion membrane?
The malate aspartate shuttle
Oxaloacetate is converted into either malate (most common) or aspartate
Malate - uses malate dehydrogenase and NADH,, with the aid of a protein it can pass the membrane and turn back into oxaloacetate
Aspartate - uses aspartate aminotransferase, no NADH and a protein
What is the second different step in gluconeogenesis?
The 7th step
Fructose-1,6-bisphosphate + H2O -> Fructose-6-phosphate + Pi
Uses Fructose bis-phosphatase
What is the third different step in gluconeogenesis?
The 9th (final) step
Glucose-6-phosphate + H2O -> Glucose + Pi
Uses glucose-6-phosphatase
What is the main regulatory point in gluconeogenesis?
Fructose bis-phosphatase - not a glycolytic intermediate but an allosteric activator
Fructose-2,6-bisphosphate activates PFK and inhibits FBPase, to maintain glycolysis when needed
How else can glucoenogenesis be regulated?
Pyrvuate kinase can be allosterically inhibited by alanine, which is converted into pyruvate via transamination
This process increases gluconeogenic flux
Insulin inhibits the transcription of the gene for PEPCK
High concentrations of cAMP promote transcription of the gene for PEPCK
Describe glycogen?
Glycogen is a polymer of a(1,4) linked D-glucose (chain length 8-14 residues) with a(1,6) linked branches every 4-8 residues
Glycogen occurs as intracellular granules (100-400 Å diameter), each contains ~ 120,000 glucose units (especially prominent in muscle and liver)
It is a storage polysaccharide, storing glucose to avoid osmotic stress
What does the breakdown of glycogen involve?
3 enzymes
Glycogen phosphorylase
Glycogen debranching enzyme
Phosphoglucose mutase
Glycogen breakdown - describe glycogen phosphorylase?
Cleaves glucose units from glycogen to yield G6P
It removes a glucose unit if it is at least 5 units away from branch point
Homodimer of 97 kDa subunits
It has a buried active site, which fits 4-5 sugars = processive enzyme
It has a covalently bound cofactor, PLP required for activity
PLP is a vitamin B6 derivative that is covalently linked via a Schiff base with a lysine residue
The phosphate of PLP acts as a general acid-base catalyst
Glycogen breakdown - what is the mechanism of glycogen phosphorylase?
- Formation of an E-P-glycogen ternary complex
- Shielded oxonium ion intermediate formation from the alpha-linked terminal glucosyl residue involving acid catalysis by Pi, as facilitated by proton transfer from PLP. The oxonium ion has the half-chair formation 3. Reaction of Pi with the oxonium ion, forms glucose-1-phosphate
Glycogen breakdown - what does glycogen phosphorylase do simply?
Catalyses glycogen phosphorolysis - bond cleavage by the substitution of a phosphate group
This yields glucose-1-phosphate
Glycogen breakdown - how is glycogen phosphorylase regulated?
Regulated both by allosteric interactions and by covalent modification (phosphorylation and dephosphorylation)
Phosphorylation at a serine or threonine is one of the most common regulatory motifs in eukaryotes
Phosphorylase a = phosphorylated state
Phosphorylase b = unphosphorylated state
Allosteric inhibitors - ATP, G6P and glucose interact with the enzyme to change between R-state = active and T-state = inactive
Glycogen breakdown - describe glycogen debranching enzyme?
Glycogen debranching enzyme is an a(1,4) transglycosidase which transfers an a(1,4) trisaccharide unit from a limit branch to the non-reducing end of a new branch
The remaining glucose unit can be hydrolysed by the same enzyme to yield glucose
This takes place due to glycogen phosphorylase only cleaves glucose units more than 5 from a branch point = the need for the removal of branch points
The enzyme has separate sites for the debranching (transferase) and hydrolysis (glucosidase) reactions
Independent active sites improves efficiency of the enzyme
Glycogen breakdown - what does glycogen debranching enzyme do simply?
Removes glycogen’s branches
This makes additional glucose residues available to glycogen phosphorylase
Glycogen breakdown - describe phosphoglucose mutase?
Phosphorylase generates G1P units which are converted to G6P by this enzyme
Major difference between this and the phosphoglycerate mutase of glycolysis is that this enzyme uses a Ser-P rather than a His-P
Glycogen breakdown - what is the mechanism of phosphoglucose mutase?
Phosphorylation of the substrate at C6-OH
Phosphorylation of the enzyme by the phosphoryl group at C1 of the substrate
Why is glycogen synthesis not a reversal of glycogen breakdown?
It is thermodynamically unfavourable without the input of energy
What does glycogen synthesis involve?
3 enzymes:
UDP-glucose pyrophosphorylase
Glycogen synthase
Glycogen branching enzyme
Glycogen synthesis - describe UDP-glucose pyrophosphorylase?
It activates glucosyl units to add to the glycogen chain
Combines G1P with uridine triphosphate to generated an “activated” compound (UDP-glucose)
UDP glucose can be added to the growing glycogen chain
This reaction is (deltaG) = 0, however coupled it is exergonic
This is a phosphoanhydride exchange reaction, this releases energy
Glycogen synthesis - describe glycogen synthase?
Extends glycogen chains
Transfers glycosyl unit of UDP-G to the C4-OH of a non-reducing end of glycogen to form an a(1,4) glycosidic bond
Highly favourable (ΔG = -13.4 kJ/mol)
Glycogen synthesis - how is glycogen synthase regulated?
Phosphorylation:
The dephospho form is more active and the phospho form is less active
Has multiple phosphorylation sites making more complex
Allosteric control
Inhibited by ATP, ADP and Pi at physiological concentrations
Almost totally inactive in vivo
Activated by G6P and allows rapid response to local needs
Glycogen sythesis - describe glycogen branching enzyme?
Amylo-(1,4®1,6)-transglycosylase
Transfers a 7 glucose unit from the end of a chain to a C6-OH group of a glucose residue on the same or another glycogen chain
Glycogen synthesis - what primes the glycogen synthesis?
Glycogenin
Acts as a glycosyltranferase, attches a glucose residue to the OH group of it’s Tyr 194
The extends the glucose chain by up to 7 donated glucose residues to form a glycogen primer
This allows glycogen synthase to bind
Glycogen synthesis - what is the ideal structure of glycogen?
2 branch points, resulting in a roughly spherical molecule that is organised in tiers (a mature glycogen has roughly 12 tiers)
If it had more branches: the ends increase however so does particle density - which limits the size of the particle
Optimum chain length = 13 residues