Carbohydrate anabolism

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Carbohydrate anabolism

• We have covered some aspects of carbohydrate
  catabolism glycolysis, PPP, citric acid cycle,
  etc. and now we turn to carbohydrate anabolic
  pathways that utilize ATP and reducing power for
  biosynthesis (ATP used to make favorable
  reactions)
• Anabolic pathways are generally reductive rather
  than oxidative
• We will use this tact in the future to cover the
  metabolism of amino acids, lipids, etc.

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Which way am I going?

• Its easiest to consider metabolic pathways as
  simple linear processes where A leads to B, B to
  C, etc.
• BUT, anabolic and catabolic pathways proceed
  simultaneously (albeit at different rates)
  producing a dynamic steady state

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Levels of organization

• Although they may share many reactions,
  biomolecules are synthesized and degraded via
  different pathways.
• Each catabolic and anabolic pathways has at least
  one unique enzymatic reaction that is essentially
  irreversible
• If not for this, flux through metabolic pathways
  would solely be due to mass action

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Unique reactions are points of control

• Like other pathways, a biosynthetic pathway is
  usually regulated at an early step that commits
  intermediates to that pathway
• Opposing (catabolic and anabolic) pathways are
  regulated in coordinated reciprocal manners

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Citric acid cycle and glyoxylate cycle

• Isocitrate conversion is the point of control
  between these two pathways
• Accumulation of citric acid cycle intermediates
  activate isocitrate dehydrogenase
• Accumulation of citric acid cycle intermediates
  inhibits isocitrate lyase

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Carbohydrate biosynthesis
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Gluconeogenesis

• A seemingly universal pathway
• reverse glycolysis Pyruvate ? glucose
• Seven of the ten reactions of gluconeogenesis are
  the reverse of glycolytic pathways
• Three glycolytic steps are essentially
  irreversible under cellular conditions
• Hexokinase, PFK-1, pyruvate kinase

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These three reactions are bypassed

• Pyruvate ? PEP
• Fructose 1,6 bisphosphate ? Fructose 6-phosphate
• Glucose 6-phosphate ? glucose

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First by-pass involves two steps

• Instead of pyruvate kinase, phosphorylation of
  pyruvate is accomplished by through intermediate
  stages involving oxaloacetate and malate
• Pyruvate is transported from cytosol to
  mitochondria (or generated from alanine within
  mitochondria via transamination)

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Pyruvate carboxylase is the first regulated step
in gluconeogenesis

• This biotin-containing enzyme was introduced via
  anaplerotic reactions
• Pyruvate carboxylase requires acetyl-CoA as a
  positive effector
• Oxaloacetate is formed through this reaction,
  which is subsequently reduced to malate via
  malate dehydrogenase and NADH

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Malate serves as a shuttle for oxaloacetate

• The resulting malate is transported to the
  cytosol via the malate a-KG transporter (from
  aspartate-malate shuttle)
• In the cytosol, malate is re-oxidized to OAA by
  cytosolic MDH
• OAA is converted to PEP by phosphoenolpyruvate
  carboxykinase

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From pyruvate to PEP
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Note the investment in activation of
intermediates through this reaction

• One ATP and one GTP used, contrasting the single
  ATP used to make PEP in glycolysis
• The CO2 added in the first reaction is released
  in the second

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Why go thru the mitochondria?

• The NADH/NAD ratio in the cytosol is 105
  times lower than in mitochondria, gluconeogenesis
  relies on NADH
• Transport of malate (reduced form of OAA)
  facilitates transport of reducing power from
  mitochondria to cytosol (subsequent generation
  of NADH by MDH) to aid in gluconeogenesis

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A second PEP biosynthetic pathway

• Lactate, instead of pyruvate, serves as a
  starting substrate in some situations (anaerobic
  muscle or erythrocytes)
• Conversion of lactate to pyruvate generates NADH
  obviating the need to export reducing power from
  mitochondria
• As a result, the PEP is generated within the
  mitochondria

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The second and third by-pass are similar

• Fructose 1,6-bisphosphate is converted to
  fructose 6-P by fructose 1,6-bisphosphatase
• Glucose 6-phosphate is converted to glucose by
  glucose 6-phosphatase
• These reactions do NOT result in ATP formation,
  instead the irreversible hydrolysis forming
  inorganic phosphate

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The cost of gluconeogenesis
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Many molecules can feed into gluconeogenesis

• This is of
• importance when we
• get to amino acid
• biosynthesis

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Reciprocity of glycolysis and gluconeogenesis

• Simultaneous operation of both glycolytic and
  gluconeogenic reactions would be wasteful if both
  reactions proceed at high rates in cells (The
  simultaneous operation of anabolic and
  catabolic pathways is a regulated process)
• Futile cycles can be engaged for physiological
  purposes such as heat energy

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Reciprocal regulation

• The first control point for regulating flux
  between these pathways is pyruvate
• Pyruvate can be converted to acetyl-CoA (pyruvate
  dehydrogenase) or to OAA (pyruvate carboxylase)
• Acetyl-CoA is a positive allosteric effector of
  pyruvate carboxylase and a negative modulator of
  pyruvate dehydrogenase

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Effects of acetyl-CoA
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A regulatory example

• When cells have enough energy, oxidative
  phosphorylation slows, NADH accumulates, inhibits
  the citric acid cycle and acetyl-CoA accumulates.
  
• This directs pyruvate to gluconeogenesis

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A second control point

• Fructose 1,6-bisphosphatase is strongly inhibited
  by AMP, while PFK-1 is activated by AMP and ADP,
  but inhibited by citrate and ATP
• Again, these opposing steps are regulated in
  coordinated and reciprocal fashion
• Also, hormonal regulation in the liver

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Hormonal regulation is mediated by fructose 2,6
bisphosphate

• fructose 2,6 bisphosphate is an allosteric effort
  for PFK-1 and fructose 1,6-bisphosphatase
• fructose 2,6 bisphosphate binds and increases
  PFK-1 affinity for fructose 6-phosphate, and
  reduces its affinity for ATP and citrate
  stimulating glycolysis
• fructose 2,6 bisphosphate inhibits fructose
  1,6-bisphosphatase

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Fructose 2,6-bisphosphateregulation
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Fructose 2,6-bisphosphate formation regulation

• Fructose 2,6 bisphosphate is generated by PFK-2
  and broken down by fructose 2,6 bisphosphatase
  (single polypeptide) note this compound is not
  a metabolic intermediate, but a regulatory
  compound

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Glucagon lowers the cellular level of fructose
2,6-bisphosphate

• Inhibits glycolysis, but stimulates
  gluconeogenesis
• Process occurs via a signal transduction pathway,
  which results in alteration of PFK-2/FBPase-2
  polypeptide