Focus on Glycolysis Flashcards
What are the 3 stages of catabolism in the presence of oxygen?
1- Acetyl-CoA production - glucose, fatty acids and amino acids all converge to make simple 2C Acetyl-CoA
2-Acetyl-CoA oxidation - citric acid cycle
3-Electron transfer and oxidative phosphorylation
Whta happens carbs, fats and proteins when digested and absorbed by chemotrophs?
-used immediately for energy via catabolic pathways
-or synthesised into glycogen (glucose store) - fuel
-or triacyclglycerols (fat store) for storgae and later catabolism - fuel
What is the major fuel in most organisms that plays a central role in metabolism and why?
D-Glucose (alpha form)
-completre oxidation of one d-glucose to CO2 and H2O after 3 stages of catabolism
-gives mega-energy - -2840kJ/mol
4 major pathways of glucose utilisation?
- storage -> glycogen, starch, sucrose
- oxidation via glycolysis -> pyruvate
- oxidation via pentose phosphate pathway
-> ribose 5-phosphate - synthesis of structural polymers -> extracellular matrix, cell wall polysaccharides
Points on glycolysis?
-1st metabolic pathway elucidated in yeast and muscle cells
-near universal pathway
-responsible for largest flux of C in cells
-role of ATP and phosphorylated compounds in cell biology discovered by research which discovered glycolysis
What happens in the Preparatory Phase in glycolysis? - 5 steps
- Glucose is phosphorylated by hexokinase to form glucose 6-phosphate
- Glucose 6-phosphate undergoes isomerisation by phosphohexose isomerase to form Fructose 6-phosphate
- Fructose 6-phosphate is phosphorylated by phosphofructokinase-1 to form Fructose 1,6 bisphosphate
- Fructose 1,6 bisphosphate is cleaved by aldose into Glyceraldehyde 3 phosphate and Dihydroxyacetone phosphate
- Glyceraldehyde 3-phosphate and Dihydroxyacteone phosphate are interconversed by triose phosphate isomerase to form Glyceraldehyde 3-phosphate
What happens in the Pay Off Phase in Glycolysis? -5 steps
- Glyceraldehyde 3- phosphate is oxidised and phosphorylated by Glyceraldehyde 3-phosphate dehydrogenase to form 1, 3-biphosphoglycerate
- 1, 3-biphosphoglycerate is involved in a substrate phosphorylation by phospho-glycerate kinase to release ATP and form 3 phosphoglycerate. ATP PRODUCTION
- 3 phosphoglycerate becomes 2 phosphoglycerate due to phosphoglycerate mutase
- A water molecule is removed from 2 phosphoglycerate by enolase to form phosphoenol pyruvate
- Phosphoenolpyruvate undergoes substrate level phosphorylation by pyruvate kinase to produce ATP and pyruvate- ATP PRODUCTION
What happens in step 1 of glycolysis and what does this involve?
Hexokinase
Glucose is phosphorylated on position 6 - ATP donates the phosphate
-Priming irreversible reaction - ATP is consumed in order to energise the system - destabilises glucose for breakdown by phosphorylating it
-Free energy becomes large and negative as ATP adds its terminal phosphate onto glucose
What are kinases?
A large family of enzymes that add phosphoryl groups to substrates - substrate level phosphorylation
What does Hexokinase require and why?
Requires Mg2+ as it shields the terminal phosphate of ATP from hydrolysis by H20 to enable nucleophilic attack by OH on C6 of glucose
Structure of hexokinase?
-Dimer and has 4 isoforms made from different genes
-Regulatory enzyme of glycolysis
-Has 2 active sites - one is regulatory site and is is phosphorylating
-Has a U-shaped clamp like structure - conformational change with clamp closing induced by binding of D-glucose
-The phosphorylation of glucose by HK traps glucose inside cells for metabolism
What happens in step 2 - Phosphohexose isomerase?
Glucose 6-phosphate isomerises to Fructose 6-phosphate
-near-equilibrium rxn - reversible
-small chnage in standard free energy
-reversible isomerisation of an aldose to a ketose
-shuffles atoms around to move carbonyl from C1 to C2 so next reaction can happen
What happens in step 3 - Phosphofructokinase-1 (PFK1)
Fructose 6-phosphate is phosphorylated tp Fructose 1,6 bisphosphate ATP dontaes the phosphate
-Primining irreversible rxn
-ATP consumed here again as in step 1 - neg free energy
-committed step for glycolysis - rate limiting step
What is PFK-1?
Majore regulatory enzyme of glycolysis
What happens in step 4 - enyzme aldolase?
Hexose is lysed (split) to form 2 trioses - Dihydroxyacetone phosphate and Glyceraldehyde 3-phosphate
-pos change in free energy - reversible rxn
-near equilibrium rxn
-C1 to C3 of Fructose 1, 6 bisphosphate makes DHAP
-C4 to C6 of F1,6-BP makes G3-P
-Lysine residue important for lysis in enzyme
What happens in step 5 - Triose Phosphate isomerase (TPI)
Isomerisation of DHAP tp G3P
-Near equilibrium rxn - interchangeable forms
-TPI pulls H off of one carbon and replaces it onto another carbon
-called a perfect enzyme
-changes molecule into useable form
What happens in step 6 - Glyceraldehyde 3-phosphate dehydrogenase?
2 glyceraldehyde 3 phosphate oxidised and phosphorylated to 1,3 bisphosphoglycerate
-oxidation transfers electrons from 2 x G3P to 2NAD to form 2NADH - energy
-prepares for energy production as v high energy of hydrolysis of 1,3 BPG
What happens in step 7 - Phosphoglycerate kinase?
-2 x 1,3 phosphoglycerate is dephosphorylated at position 1 to yield 2 x 3-phosphoglycerate and 2 ATP
-Substrate level phosphorylation
What role does the induced fit motion in phosphoglycerate kinase play?
-Same as hexokinase
-Closes around rxn, protecting it from interfering with water molecules so ATP is not hydrolysed immediately
-2 lobes - upper lobe binds ADP and lower pocket binds 1,3BPG - hinges closes to perform transfer of phosphate
What happens in step 8 - Phosphoglucomutase?
2 x 3 phosphoglycerate isomerises to 2 x 2 phosphoglycerate
-the phosphate group transfers onto enzyme active site before it moves to position 2
-in presence of magnesium
-creates a more amenable susbtrate for rxn 9
What happens in step 9 - Enolase?
-2 x 3 phosphoglycerate dehydrated to yield 2 x phosphoenolpyruvate and 2 x H2O
-this dehydration places the phosphate in an uncomfortable position making it easy to remove to form ATP in next rxn - the new double bond is in an awkward place
-Phosphoenolpyruvate is a high energy molecule
What happens in step 10 - Pyruvate kinase?
-2 x phosphoenolpyruvate is dephosphorylated to yield 2 x pyruvate and 2 ATP
-2nd substrate level phosphorylation
-regulatory enzyme
Structure of pyruvate kinase?
-An allosteric enzyme
-4 flexible subunits arranged in a diamound shape
-Senses levels of energy metabolites and turns on/off according to energy needs
-these allosteric modulators chnage shape of enzyme active <-> inactive
What is the total energy gained from glycolysis and what is this pathway?
2ATP and 2NADH
Exergonic pathway
(2 ATP invested and 4 produced)
Why is this pathway unusual?
As things not usually broken down in the cytosol
Why is the foirmation of glucose -6 phosphate not a rate limiting step (first step)?
As can be used in the cell anways in that form
-second phosphorylation is rate limiting step- the PFK-1
What is the 3 possible fates of pyruvate?
In presence of oxygen - becomes 2 Acetyl CoA and enters citric acid cycle
Absence of oxygen - ethanol and CO2 (in yeast) or Lactate (in muscles)
Fate of Pyruvate in presence of oxygen?
- 2 pyruvate decarboxylated to 2 acetyl-CoA (2C) and 2CO2
- 2 x acetyl-CoA enters citric acid cycle to form citrate oxidised with electron transfer to NAD and FAD - committed step
- NADH and FADH2 transfer electrons to respiratory chain in to create ATP energy
-All happens in mitochondria
How muc total energy is porduced from oxidation of 1 glucose aerobicallly with the 3 stages?
2850kJ energy
In anerobic conditions what enzymes produce lactate and ethanol from pyruvate?
Pyruvate - lactate dehydrogenase - lactate
Pyruvate - pyruvate decarboxylase - acetaldehyde - alcohol dehydrogenase - ethanol
Why cant vertebrates convert pyruvate to ethanol?
Pyruvate decarboxylase is absent in vertebrates and lactic acid bacteria
What essential regeneration takes place in anaerobic conditions?
Regeneration of NAD+ which keeps stepp 6 glycolysis supplied with NAD+ allows continuous glycolysis to occur in absence of O2
When does lactate production happen?
- Wwide range of organisms in anaerobic niches produce lactate from glucose for energy fermentation of sugars to lactate by m/o used in cheese yogurt production
- erythrocytes, retina, brain cells produce lactate even even O2 present - responsible for atleats 10% overall glucose breakdown
- Very active skeletal muscle uses anaerobic pyruvate - lactate conversion when O2 cant get to muscle fast enough - anaerobic respiration
- Cancer cells and immune cells can have very active anaerobic glucose catabolism even in presence of O2 (Warburg effect) for fast proliferation
Whta happens when anaerobic respiration takes place in muscles?
-O2 cant get to muscle fast enough
-Muscle switches to anaerobic glycolysis
-Breaks down muscle glycogen stores -> ´glucose -> pyruvate -> lactate for energy
-Lactate produced blood -> liver
- Lactate synthesises glucose in liver
-glucose goes from liver -> blood -> muscle replenishes glycogen
What glycolytic enzymes can occur in cancer cells?
-Increased levels of glycolytic enzymes
-Diff isoforms of glycolytic enzymes can occur including Pyruvate kinase PKM2 rther than PKM1
-PKM2 has reduced catalytic activity compared to PKM1 - allows cancer cells to keep using anaerobic glycolysis for energy and build up glycolytic intermediates that feed into synthetic pathways promoting proliferation
