Regulation of glycolysis
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Transcript of Regulation of glycolysis
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Regulation of glycolysis• Flux through biochemical pathways depends on
the activities of enzymes within the pathway• For some steps, the reactions are at or near
equilibrium in the cell• The enzyme activity is sufficiently high that
substrate equilibrates with product as fast as substrate is supplied.
• Flux is thus substrate limited
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Flux through a multi-step pathway
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Glycolysis has a bottleneck at the phosphofructokinase catalyzed step
• The rate of fructose 6 phosphate to fructose 1,6 bisphosphate is limited by PFK-1 activity
• Can produce as much fructose 6 phosphate as you want, but still won’t push glycolysis
• PFK-1 acts as a valve• This is an enzyme-limited reaction, and also
the rate-limiting step in glycolysis
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Glycolytic enzyme and metabolite balances
• Table 15-2
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Trademarks of rate-limiting steps
• Rate-limiting steps are very exergonic reactions, essentially irreversible under cellular conditions
• Typically, the enzymes that catalyze these reactions are under allosteric control
• Often, these enzymes are situated at critical branch points in metabolism
• For glycolysis, the first committed step is the PFK-1 mediated reaction
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PFK-1 is under complex allosteric regulation
• Glucose-6-phosphate can flow into glycolysis or other pathways, PFK-1 commits substrate to glycolysis. – PFK-1 is first unique step, not hexokinase.
• Several allosteric sites on PFK-1– ATP is not only a substrate but a product of the metabolic
pathway in question and inhibits PFK-1 by lowering affinity for fructose-6-P
– ATP effect countered by ADP and AMP– Citrate, a key TCA cycle intermediate, enhances ATP effect.
High [citrate], more inhibition– PFK-1 is inhibited by protons, thus senstive to pH change– Fructose 2,6 bisphosphate activates the enzyme
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Regulation of PFK-1
• Fig 15-18
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Fructose 2,6 bisphosphate?• This metabolite has an important role in switching
glycolysis and gluconeogenesis (chapter 20)• Fructose 2,6 bisphosphate is synthesized from
fructose-6-phosphate by phosphofructokinase-2 (PFK-2)
• PFK-2 is a unique enzyme, because this polypeptide also acts as fructose bisphosphatase 2 (FBPase2) which converts Fructose 2,6 bisphosphate to fructose-6-phosphate
• A bifunctional enzyme
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Hexokinase is a site for regulation in glycolysis
• Catalyzes the entry of free glucose into glycolysis
• When PFK-1 is inhibited both Fructose-6-phosphate and glucose 6-P build up. Glucose-6-phosphate inhibits hexokinase.
• Many distinct forms of hexokinase, which all convert glucose to glucose-6-phosphate.
• These multiple forms are called isozymes
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Why isozymes?
• Isozymes resulting from gene duplication events allow evolution to tune the metabolic potential of cells – Different metabolic patterns in different tissues– Different locations and metabolic roles for
isozymes in th same cell– Different stages of development– Different responses of isozymes to allosteric
modulators
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For instance,
• Hexokinase expressed in liver has distinct properties from the enzyme expressed in muscles– Higher Km for glucose– Inhibited by Fructose-6-phosphate, not glucose-
6-phosphate– Inhibition is mediated by a regulatory protein
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A last regulatory step – Pyruvate kinase
• Again, multiple isoforms or isozymes, which respond to distinct metabolic cues
• Pyruvate kinase found in muscle is activated by Fructose 1,6 bisphosphate (pulling intermediates through the pathway)
• Inhibited by ATP and alanine (feedback inhibition; alanine serves as a monitor for biosynthetic precursors)
• Also under hormonal control - glucagon
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Pentose phosphate pathway (PPP)
• Although most glucose is used to generate energy via glycolysis and TCA cycle, cells also need precursors for important biomolecules such as nucleic acids.
• PPP generates reducing power (NADPH) and pentoses
• Also known as Phosphogluconate Pathway, or Hexose Monophosphate Shunt
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Initial reactions generate NADPH
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As does a subsequent step…
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Ketose is isomerized to aldose or it’s epimer
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PPP
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The pathway can end here
• The ribose 5 phosphate can serve as a precursor for nucleic acid biosynthesis
• This pathway has also generated reducing power in the form of NADPH
• However, there is a cycle that allows the ribose-5-phosphate to be recycled in cells looking for more NADPH – referred to as the nonoxidative reactions of PPP
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The first step in the nonoxidative reactions
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Transketolase
• Is a TPP dependent enzyme
• Catalyzes the transfer of a two carbon fragment from xylulose 5-phosphate to ribose 5-phosphate forming a seven carbon product and glyceraldehyde 3-phosphate
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Transaldolase then generates Fructose 6-phosphate
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Completing the cycle
• Fructose 6-phosphate can be isomerized to glucose 6-phosphate and re-initiate PPP
• Meanwhile, transketolase uses erythrose 4-phosphate and xylulose 5-phosphate to produce fructose 6-phosphate (for isomerization) and glyceraldehyde 3-phosphate (for reverse glycolysis to glucose 6-phosphate)
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Oxidative and non-oxidative PPP reactions
• Fig 15-21
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The outcome of PPP is 6 carbon carbohydrates
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• Depending on relative needs of a cell for ribose-5-phosphate, NADPH, and ATP, the Pentose Phosphate Pathway can operate in various modes, to maximize different products
• There are three major scenarios:
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Make NADPH and Ribose-5-phosphate (oxidative reactions)
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Crank out NADPH (cycle with non-oxidative reactions)
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Feed products back to glycolysis to generate NADPH and ATP
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How is PPP regulated?
• Glucose 6-phosphate dehydrogenase catalyzes the first committed step, hence it is the point of regulation.
• Glucose 6-phosphate dehydrogenase activity responds to levels of NADP+
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Other glucose energy-generating metabolic pathways
• Phosphoketolase pathway– The phosphoketolase pathway is distinguished by the key
cleavage enzyme, phosphoketolase, which cleaves pentose phosphate into glyceraldehyde-3-phosphate and acetyl phosphate. (|| pentose phosphate pathway)
• Entner-Doudoroff pathway– The E-D pathway yields 2 pyruvate from glucose (same as
glycolysis) but like the phosphoketolase pathway, oxidation occurs before the cleavage, and the net energy yield per mole of glucose utilized is one mole of ATP.
ETC.