Glycogenesis and Glycogenolysis

Glycogen

• Polysaccharides tailored for storing and releasing energy on demand

  • Branched polymer of glucopyranose rings, well-suited for storage function with many accessible side chain ends and chemical stability through glycosidic linkages that prevent linearization

• Glc units are joined by O-glycosidic linkage

  • Free OH of the anomeric C in the ring has been converted into an ether linkage

• Notation

  • Glc alpha(1>d4)Glc

    • First carbon uses its anomeric carbon with an alpha-stereochemistry to form a O-glycosidic bond on the C4 on the second glucose

    • D means the second glucose is in D configuration

• Every glucose unit in glycogen is prevented from linearization,  stable storage molecule

• All sugars are D-Glucose, with main-chain linkages with alpha1>4 and branch points uses alpha1>6

 Lactose

• Galactose + glucose

  • Has reducing-sugar chemistry and would not be a wise choice for energy storage

  • Non-reducing end: anomeric carbon has acetal (aldose) or ketal (ketose) (has 2 ethers and an H)

  • Reducing end: anomeric carbon has hemi-acetal (aldoses) or hemiketal (ketose) (one ether and free OH and an H)

  • Deprotonating OH on the anomeric C enables linearization

  • Gal(beta1>4)Gluc (beta)

    • Galactose is fixed as beta

    • Glucose is not fixed as alpha or beta

Glycogenin

• Reducing end has the C1 taking part  in an O-glycosidic linkage to a protein: glycogenin

  • Cannot linearize and glycogen doesn't exhibit reducing sugar chemistry

  • Binds to the first glucose to protect the reducing end

• Glycogen has many chain ends far from glycogenin but all of them are non-reducing

  • Extensive branching provides many sites for glucose addition and removal

Pom-Pom shape

 protects reducing ends and exposes non-reducing ends

• Unique reducing end has C1 in O-glycosidic linkage

  • It cannot actually linearize and glycogen does not exhibit reducing-sugar chemistry

• Glycogenin at the centre of the pom pom and branches out

  • Protects the reducing end

A chains

• Terminal chains with no further branches emanating from them

• Accessible for glycogen processing enzymes (synthase and phosphatase)

B chains

Internal highly branched chains, serve as structural backbone and are not accessible to glycogen processing enzymes

Pom Pom shape

• Many free non-reducing ends distal from glycogen. Enzymes removes Glucose from the non-reducing ends

• Extensive branching allow many glucose molecules to be release simultaneously

Glycogen linkages

• Made of alpha units linked from axial to equatorial positions

  • C1 is axial down, C4 is down equatorial

• Forms a curved polymer that keeps branches compact

• Ideal for energy storage and utilization

Cellulose

• Straight

• Made of beta-glucose units, links equatorial to equatorial

• Forms a straight polymer that is unbranched

• Packs in parallel for fibrous structure

• Ideal for cell wall

glycogenesis vs glycogenolysis

• Anabolic pathway to build glycogen from glucose

• G6P is converted to G1P

• Forms activated precursor UDP-glucose used to add glucose units to glycogen

release G1P from glycogen's non-reducing ends, G1P is converted to G6P which feeds into glycolysis

 

Glycogenesis Step 1

• Phosphoglucomutase-catalyzed the reversible conversion of G6P to G1P

• Free energy is 1.1 kJ/mol, near 0, reaction is near equilibrium, allows it to proceeds in either direction depending on concentration of substrate or product

Purpose

• Essential in glycogen synthesis as G1P is needed to form UDP-Glucose

• Allows cells to efficiency redirect glucose from glycolysis to be stores as glycogen

Mechanism

• Phospho-serine intermediate donates a phosphate to 1-posiotn of C6P, forms glucose-1,6-bisphosphate

• Flips over to transfer C6 phosphate back to enzyme, releases G1P

Glycogenesis step 2

Glc1-P + UTP > UDP-G + PPi

• UDP-glucose pyro phosphorylase converts G1P to UDP-glucose, activated donor for glycogen synthesis

• UDP-glucose pyro phosphorylase catalyzes the conversion of G1P and UTP into UDP-glucose, and pyrophosphate

• Reaction has a free energy near 0 so it is close to equilibrium, in vivo it proceeds forward because Ppi is rapidly hydrolyzed to 2 Pi by pyrophosphate, makes free energy highly negative

  • Net free energy is still highly negative

Purpose

• Key reaction forms activated sugar donor (electrophile) required glycogen synthesis

• The high energy bond between UDP and glucose and provides energy for the following reaction

Mechanism

• Enzyme position G1P and UTP in the active site for a nucleophilic attack of the C1-phosphate on the electrophilic alpha-phosphate of UTP, forms UDP-glucose and release Ppi

Glycogenesis Step 3: Priming

UDP-glucose + glycogenin (glucosyltransferase activity) releases UDP and glucose is linked to tyrosine. Chain-extending activity occurs, with UDP leaving as a product in each addition

Glycogenin uses UDP-glucose to add the first few glucose residues

• Glycogenin both enzyme and primer to initiate glycogen synthesis

• Catalyzes glucosyltransfer of glucose from UDP to Tyr 194, creates the first glycosidic bond and elongates a short alpha 1>4 linked glucose chain

• Standard Free energy of glycosyltrasnfer is near 0, in the cell the reaction proceeds forward because UDP is rapidly hydrolyze to UMP and Pi to drive the process

Purpose

• Priming is essential as glycogen synthase cannot state from scratch, requires a pre-existing chain to extend

• Glycogenin acts as protect by anchoring the reducing end, prevent linearization and exposure of the reactive aldehyde

Mechanism

• Active-site tyrosine hydroxyl as nucleophile to attack the anomeric carbon of UDP glucose, releases UDP and establish the first glucose-protein linkage

 

Glycogenesis Step 4: Glycogen synthase

UDP-glucose (glycogen synthase) > extending chain

Glycogen synthase catalyzes elongation of glycogen

• Transfer glucose from UDP-Glc to the nonreducing end of the glycogenin primer

• Forming alpha 1>4 glycosidic bond

Standard free energy, cellular UDP removal drive it forward

Purpose

• Commits activated glucose into glycogen for energy storage

• Glycogen synthase is progress, it can go multiple rounds of catalysis without release the product

Mechanism

• Terminal C4 hydroxyl of the glycogen chain performs a nucleophilic attack on the anomeric carbon of UDP-glucose

• Releases UDP and extends the chain with high specificity for alpha 1>4 linkages

Glycogenesis step 5

Branching enzymes transfers glucose from UDP-Glucose to form alpha1>6 branch points

Purpose

• Branches are essential for glycogen to form the pom pom shape to have accessible glycogen ends

• Branching cannot be accomplished by glycogen synthase which only adds alpha 1>4 linkage

Branching enzymes

• Alpha 1,4 > alpha 1,6-transglycosylase cleaves a terminal fragment of s6 to 7 glucose residues form an existing alpha 1>4 glycogen and transfer it to a C6 hydroxyl group of glucose residue located in the interior of the same or another glycogen chain

• Forms a new alpha 1>6 glycosidic linkage and makes a branch point

Mechanism

• Glycosyltransfer, simple hydrolysis to break the alpha 1>4 glycosidic bond followed by a condensation from the new alpha 1>6 glycosidic

• Standard free energy is near 0, reaction is driven forward by high availability of linear glycogen chains (reaction quotient effect)

Glycogenolysis Step 1

Glycogen phosphorylase catalyzes the phosphorolytic cleaves of alpha1>4 glycosidic bonds at the nonreducing ends of glycogen produces G1P, Pi is a reactant

• Pi releases G1P and is rearranged to G6P

• Pi acts a nucleophiilc to split glycosidic bond (O-glycosidic bond is low-energy)

• WITHOU ATP being consumed and glucose gets phosphate tag on anomeric C1

  • Free energy is near 0, in cell it is -25 kJ/mol because of high Pi and rapid G1P utilization

Irreversible commited step in glycogenolysis that glycogenesis needs a bypass

Purpose

• Mobilizes G1P from glycogen, rapid conversion to G6P for glycolysis or PPP without consuming ATP

Mechanism

• Employs pyridoxal phosphate PLP cofactor to stabilize the transition state and facilitate proton transfer during cleavage

Glycogenolysis Step 2: Whacking down the branches, debranching enzyle resolves glycogen branch points

• Glycogen debranching enzymes resolves the alpha1>6 branch points that glycogen phosphorylase cannot cleave, allows complete mobilization of glucose from glycogen

Purpose

• Allow maximal G1P release, prevents glycogenolysis from stopping at branches residues

Mechanisms

• 1 enzyme, 2 active sites, 2 jobs

  • Glycosyltransferase activity shifts trisaccharide unit from the branch to a nearby chain and leaves an alpha 1,6-gluc at the branch

  • Glucosidase activity hydrolyzes the remaining Glucose

    • Exploit favorable entropy (breaking down sugar) and enthalpy (release 1>6 strain link)

Glycogenolysis 3

• Phosphoglucomutase catalyzes the conversion of G1P to G6P

  • Moves the phosphate tag to C6 for glycolysis entry

Endogenous glycogen enters glycolysis at G6P where dietary glycogen enter as glucose

1. Glc from endogenous glycogen are harvested/targged

   1. No use for hexose and ATP

   2. From diet it requires ATP at hexokinase step

      1. Alpha-amylase is required to hydrolyze glycogen so monosaccharides can cross from the gut to blood

Anabolic NTP cost of gluconeogenesis: Lactate to glc

2 Lactate + 6 NTP > Glc + 6Pi + 6NDP

Profit from catabolic NTP generation from glycolysis

Glc + 2Pi + 2ADP > 2 lactate + 2ATP

Anabolic NTP cost of lactate to glycogen extension

2 lactate + glycogen(n) + 7NTP > glycogen (extended) + 7 Pi + 7NDP

Higher NTP cost for anabolism than catabolic profit

Glycogen + 3Pi + 3 ADP > glycogen + 2 lactate + 3 ATP

Phosphorylase control catabolism, synthase controls anabolism

• Phosphorylase commits glucose to metabolism, very negative free energy, in glycogen synthesis the step is bypassed

  • Glc is activated to UDP-Glc by UTP glucose pyrophosphorylase and synthase elongates the chain

    • Anabolism sacrifices phosphoanhydrides to bypass irreversible catabolic steps

Regulation of Glycogen Phosphorylase

• Activated by metabolites and hormones that signal an energy need

• Inhibited by metabolites that signal energy surplus (ATP + G6P)

Regulation of Glycogen synthase

• Activated by signals of energy surplus

  • G6P, insulin

• Inactivated by signals of energy need

  • Epinephrine, glucagon

Glycogen phosphorylase has an on switch

Glycogen phosphorylase has an on switch

• 2 conformation

• Phosphorylase b is the inactive conformation

• Phosphorylase a is the active conformation

  • Stabilize by adding a phosphate tag by a protein kinase

  • Can be removed by a protein phosphatase

• Phosphorylase b kinase is activated by epinephrine, Ca2+ and AMP in muscle and glucagon in liver for times of need

  • Slower creation of glycogen (need glucose for energy)

 

Phosphorylase b kinase

• Protein kinase adds phosphoryl groups to glycogen phosphorylase b to activate it

  • Adds phosphate tag to inactivate phosphorylase b and convert it to active

    • Phosphorylase b makes 100x molecules, phosphorylase a makes 1000 molecules

• Signal amplification

  • Each kinase enzyme activates multiple copies of its target

  • Each active phosphorylase a enzyme cleave multiple glucose units

Extracellular hormones produces intracellular signals

• Hormones like glucagon and epinephrine bind to receptor cells on the cell surface, triggers cascade of conformational switching in intracellular enzymes

  • Glucagon/epinephrine increase cAMP and binds to PKA

  • Increase activity to phosphorylate and activate phosphorylase b kinase causing the downstream affect

Regulation of glycogenesis

• Glycogen synthase has an OFF switch

  • Synthase activated when dephosphorylated and inactive when phosphorylated, reciprocal regulation with glycogen phosphorylase

  • Ensures coordinated control of glycogen synthesis and breakdown

• Active: GSa, dephosphorylated

• Inactive: GSb, phosphorylated

• Hormones/metabolites modulate GS3K kinase and protein phosphatase 1 which act reciprocally to toggle the enzyme

• Active: Dephosphorylated Gsa

  • Times of plenty signal PP1 to dephosphorylate GSb (insulin, G6P, G)

    • Insulin inhibits GSK3 keeping glycogen synthase active (GS3K will phosphorylate GS)

  Inactive: Phosphorylated GSb

  • Glucagon/epinephrine block PP1 catalyzed dephosphorylation of glycogen synthase a

   

  GS is regulated by reciprocal phosphorylation to ensure glycogen synthesis occurs when energy and nutrients are abundant