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What shape is the Glycolysis pathway?
Linear
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Glycolysis (general facts)
- series of biochemical rxns that convers 1 glucose molecule to 2 molecules of pyruvate with release of usable E in form of 2 ATP molecules
- an anaerobic process in the cytoplasm
- on it's own an inefficient metabolic pathway for E generation
- pyruvate can be futher fermented to lactate or ethanol
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one molecule of glucose = ?
one molecules of glucose = (2) 3-C molecules (of pyruvate) and 2 molecules of ATP
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Major stages in glycolysis
- I- conversion of glucose to fructose-1,6-bisphosphate (2 ATP used)
- II- generation of 2 inter-convertible 3-carbon molecules, dihydroxyacetone phosphate (DHAP) and glyceraldehyde 3-phosphate (GAP)
- III- conversion of DHAP and GAP to pyruvate (each 3-carbon molecule oxidised to produce 2 ATP molecules and 1 NADH)
- Net gain of 2 ATP and 2NADH molecules
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Key regulators in glycolysis
- Phosphofructokinase
- pyruvate kinase
- pyruvate dehydrogenase
- (regulate the rate of glycolysis process)
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Glycolysis (Stage I)
- Hexokinase phosphorylates glucose (G) to G-6P (1 ATP used)
- Phosphoglucose isomerase isomerises G-6P to fructose (F)-6P (6 to 5 member ring conversion)
- Phosphofructokinase (PFK) phosphorylates F-6P to fructose 1,6-bisphosphate (F-1,6BP) (1 further ATP used)
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Glycolysis (Stage II)
- F-1,6BP is split into isomers, DHAP and GAP (interchangable), by aldolase (highly reversible)
- DHAP converted reversibly to GAP by triose phosphate isomerase, hence 1 glucose gives 2 GAP molecules
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Glycolysis (stage III)
- GAP dehydrogenase (GAPDH) catalyses oxidation of GAP to 1,3-bisphoshoglycerate (1,3-BPG) with production of NADH and H+ (reduced)
- 1,3-BPG has high phosphoryl transfer potential. Phosphoglycerate kinase transfers phosphate group to ADP to form 1 ATP molecule
- 3-PG is converted to 2-PG by Phosphoglycerate mutase
- 2-PG is dehydrated to phosphoenol-pyruvate (PEP) by enolase
- high phosphoryl transfer potential of PEP catalysed to pyruvate by pyruvate kinase (PK) with formation of 1 ATP
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Glycolysis net yield
- Glucose to pyruvate net yield: Glucose + 2Pi + 2 ADP + 2 NAD+
- --> 2 pyruvate + 2 ATP + 2 NADH + 2H+ + 2H2O
- redox balance and outcome of pyruvate
- reduction of pyruvate and acetaldehyde by NADH to lactate (LDH) and ethanol (ADH) respectively
- (krebs cycle)- regeneration of NAD+
- pyruvate transferred to mitochondra, decarboxylated and oxidised to form acetyl CoA
- Pyruvate + NAD+ + CoA--> acetyl CoA + CO2 and NADH (pyruvate dehydrogenase complex)
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Cori cycle
- in skeletal muscle undergoing intense contraction and in RBC (no mitochondria), pyruvate and NADH accumulate. NAD+ is regenerated by reduction of pyruvate to lactate (LDH)
- lactate diffuses into circulation to liver. where it is oxidised back to pyruvate and subsequently converted into glucose to be fed back to muscle (allowing glycolysis to cont. w/o O2)
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Fructose enters glycolysis pathway at intermediate stages...
- Fructose is converted by fructokinase to fructose 1-phosphate (F-1P). F-1P split into DHAP and GAP by another aldolase, F-1P aldolase. REQUIRES ATP (for their breakdown)
- alternatively fructose converted (with less affinity than glucose) by hexokinase to F-6P. requires ATP
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Galactose enters glycolysis pathway at intermediate stages...
- galactose is converted to glucose-6P in 4 steps
- (galactokinase, galactose 1- phosphate uridyl transferase, epimerase, phosphoglucomutase)
- Galactose + ATP---> glucose 6-phosphate + ADP + H+
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Milk intolerance
- insufficient lactase in the gut to cope with milk intake
- lactase breaks down lactose to glucose and galactose
- post- weaning lactase levels down to 5 to 10% of birth
- microbial breakdown of lactose to lactic acid, methane and hydrogen. undigested lactose and lactic acid osmotically draw fluid into gut lumen, hence diarrhoea
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Regulatory checkpoints of glycolysis
- irreversible reactions!! of (regulatory checkpoints):
- hexokinase
(G --> G-6P) - Phosphofructokinase (PFK) (F-6P --> F-1,6BP)
- Pyruvate kinase (PK) (PEP --> pyruvate)
- the corresponding genes are regulated at the level of transcription and translation
- their enzymatic activities are regulated by allosteric effects or by covalent modifications
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Regulation of PFK activity
- PFK is a tetramer, each subunit with a catalytic and an allosteric site
- INHIBITORS of PFK:
- -allosteric ATP binding to PFK reduces affinity of PFK to F-6P
- -rise in [H+] or fall in pH inhibits PFK (as in lactic acid build up)
- -Citrate, an early intermediate of the citric acid cycle, enhances the inhibitory effect of ATP
- -Low glucose level
- ACTIVATORS OF PFK:
- -AMP reverses the allosteric inhibition of ATP
- -High Glucose level
- -Allosteric F-2,6BP (produced in the liver) binding stimulates PFK (NOT F-1,6 BP)
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Allosteric activator: F-2,6 BP
- Glucose high: PFK2 (dual kinase and phosphatase) converts F-6P to F-2,6BP, which in turn stimulates PFK (note PFK2 does not equal PFK)
- Glucose Low: --> glucagon increases --> protein kinase A --> phosphorylation of PFK2 --> activation of it's phosphatase domain --> reduced F-2,6BP --> reduced PFK activity (in liver)
- (glucagon promotes glycogenolysis and gluconeogenesis)
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Hexokinase
- Hexokinase (G --> G-6P) is inhibited by G-6P
- inhibition of PFK (F-6P --> F-1,6BP) will eventually lead to build up of G-6P. hence inhibition of PFK will also inhibit hexokinase
- Role of isozyme glucokinase (activitaed w/ glucose levels are very high)(Gk, lower affinity for glucose and not inhibited by G-6P):
- when glucose high, Gk provides G-6P for glycogen synthesis (G-6P can also be oxidised by pentose phosphate pathway to generate NADPH)
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Pyruvate Kinase (PK)
- PK (PEP --> pyruvate + ATP), a tetramer, encoded by different genes to give rise to L-type (liver isoform) and M=-type (muscle and brain isoform)
- Activators of PK:
- -F-1,6BP (product of PFK) activates both PK isoforms
- -insulin signalling leads to dephosphorylation (activation) of PK
- Inhibitors of PK:
- -ATP allosteric inhibition of both PK isoforms
- -alanine (made from pyruvate) allosteric inhibition (abundance of building blocks)
- -glucogon signalling leads to phosphorylation (inactivation) of L isoform of PK
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Regulation of rate of glycolysis at level of Pyruvate kinase (liver isoform)
- high glucose (insulin) phosphorylated pyruvate kinase (less active)
- dephosphorylated pytuvate kinase (active)->Phosphoenolpyruvate + ADP + H+-->Pyruvate + ATP
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Pyruvate kinase deficiency
- an inherited autosomal disorder
- reduced survival of red blood cells (no mitochondria and nucleus) from deficiency in ATP
- loss of biconcave shape and haemolytic anemia
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Pentose phosphate pathway (PPP)
- CYTOSOL
- primarily anabolic. utilizes 6 carbons of glucose --> 5 carbon sugars and reducing equivalents (NADPH)
- however, does oxidise glucose and under some conditions can completely oxidize glucose to CO2 and H2O
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PPP relationship to glycolysis
PPP pathway branches from glycolysis at level of G-6-P
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primary functions of PPP
- to generate reducing equivalents, in the form of NADPH, for reductive biosynthesis
- to provide the cell with ribose-5-phosphate (R5P) for the synthesis of the nucleotides and nucleic acid
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NADPH for reductive biosynthesis
- in nucleated cells w/active lipid biosynthesis (eg lactating mammary glands, adrenal cortex and liver)
- NADPH is used in redox rxns required for biosynthesis of fatty acids, cholesterol, steroid hormones, and bile salts
- IN LIVER: NADPH used for hydroxylation rxns involved in detoxification and excretion of drugs
- IN RBCs: NADPH used in reduction of glutathione
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PPP secondary function
metabolize dietary pentose sugars (5 carbon atoms) derived from the digestion of nucleic acids as well as to rearrange the carbon skeletons of dietary carbohydrates into glycolytic/gluconeogenic intermediates
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PPP shunt
- when pentoses not needed for biosynthetic rxns, pentose phosphate intermediates are cycled back into mainstream of glycolysis by conversion in f-6P and glyceraldehyde-3-phosphate
- (shunted back so nothing is wasted)
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