BIOC C NOTE

1. General Metabolism Concepts

Metabolism - Concepts

  • Metabolism is the SUM of chemical transformations
  • Metabolic pathways involve enzymes
  • Catabolism = degradation of biomolecules
  • Anabolism = synthesis of biomolecules

Types of Pathways

  • Converging catabolism
  • Cyclic pathway
  • Diverging anabolism

Concepts Review

  1. Metabolism is the sum of reactions
  2. Catabolism is the breakdown of molecules
  3. Anabolism is the synthesis (biosynthesis) of molecules
  4. Rate-limiting steps determine the overall speed of a pathway
  5. Rate- limiting steps represents points of regulation, and are exergonic
  6. Being strongly exergonic = reactions essentially irreversible

2. Rate limiting Enzymes

Rate - Limiting Steps

  • Rates of biochemical pathways depend on the activities of enzymes that catalyze each step
  • Reactions that catalyze one or more enzymes in any pathway will be limiting
  • Rate-limiting steps are often exergonic and irreversible under cellular conditions
    • ENZYMES catalyzing exergonic rate-limiting steps are targets of METABOLIC REGULATION

3. Glycolysis and regulation of:

1. Hexokinase (step 1)

  • Glucose → G-6-Phosphate (using enzyme Hexokinase, and ATP → ADP)

    Hexokinase is allosterically inhibited by its product

    • Hexokinase I is expressed in muscle (max activity)
    • if G6P increases, enzyme is inhibited (-ve feedback)
    • Hexokinase IV (glucokinase) is expressed in the liver
    • high blood glucose relieves inhibition by glucokinase regulatory protein
    • lower affinity for glucose
    • inhibited by fructose 6-p (glucokinase regulatory protein)
      • increase F6-P = binds to regulatory protein
    • IV = HIGH BLOOD GLUCOSE = IN CYTOSOL
    • I = LOW BLOOD GLUCOSE = SEQUESTERED INTO THE NUCLEUS

    Hexokinase IV has a relatively poor K0.5

    • enzyme I do not increase rate when glucose is higher than optimal
    • IV has higher K0.5 (10 mM) compared to 0.2 mM in the I enzyme

    Regulation

    • High fructose-6-p = glucokinase regulatory protein sequesters hexokinase IV in the nucleus
    • F6P binds to Regulatory protein causing a conformational change
    • Regulatory protein binds to Hexokinase IV causing another conformational change (sprouts NLS)
    • High glucose weakens the enzyme/regulator interaction (encourages cytosolic localization)

2. Phosphofuktokinase-1 (step 3)

  • Fructose 6 phosphate + ATP → Fructose 1,6, bisphosphate + ADP (using citrate, and Fructose 2,6, bisphosphate)

  • phosphorylation by PFK-1 commits fructose-6-phosphate to glycolysis (in equilibrium with G6P)

    PFK-1 Allostery

    • ATP = negative heterotropic modulator (lowers affinity for fructose 6-P)
    • ADP and AMP relieve inhibition by ATP
    • Citrate increases inhibition by ATP
    • Fructose 2,6, bisphosphate = strong activator

    Regulation of PFK-1 by ATP

    • High ATP = reduce affinity of PFK-1 for f6p
    • low ATP = high F6P affinity allows PFK-1 to be more active

3. Pyruvate Kinase (step 10)

  • Phosphoenolpyruvate → (2) Pyruvate (2ADP → 2ATP)

    Pyruvate Kinase inhibition

    • High ATP allosterically inhibits PK, decreasing its affinity for PEP
    • Acetyl-CoA and long-chain fatty acids also inhibit PK
    • when plentiful, so is ATP
    • ALANINE (- allosteric modulator)
    • F 1,6-BP accumulation (+ allosteric modulator)

4. Gluconeogenesis and regulation of:

1. Step 1

  • Bypass for step 10 of glycolysis (pyruvate → phosphoenolpyruvate/PEP)
  • Gluconeogenesis consumes NADH
  • Liver produces NADH in the mitochondria during gluconeogenesis (degrading fatty acids)
  • NADH = not transported to the cytosol
  • mitochondrial malate dehydrogenase consumes NADH, cytosolic malate dehydrogenase produces NADH
  • Lactate dehydrogenase produces cytosolic NADH (feedback for gluconeogenesis)
  • Liver PEP carboxylase produces PEP directly, and extra NADH is used (not needed in cytosol)

2. Step 10

  • G-6-Phosphatase catalyzes the dephosphorylation of G-6-P
  • Expressed in a few tissues (liver, kidney, small intestine) = gluconeogenic tissues

3. Step 8

  • Fructose 1,6 bisphosphatase converts F 1,6, Bis phosphate to F 6-P (phosphatase)

  • AMP(-)

  • F 2,6-BP (-)

    STEP 3

  • F 2,6 - BP (+)

  • AMP (+)

  • ADP (+)

  • ATP (-)

Citrate (-)

F26PB = positive modulator for Glycolysis, negative modulator for gluconeogenesis

Positive regulator for PFK-1 and negative modulator for FBPase-1

  • PFK-1 low affinity for fructose-6-P in the absence of F2, 6BP

  • F2, 6BP has high affinity for fructose-6-P

  • FBP-1 = high affinity for f-1,6-BP in absence of F26PB

  • Affinity for F16PB decreases in the presence of F26PB

F 2,6 PB = controlled by two opposing enzyme activities: PRK-1 and Fructose 2,6 - bisphosphatase

  • PFK-2/FBPase-2 = BIFUNCTIONAL enzyme
    • phosphorylation by PKA = activates FBPase 2 and INACTIVATES PFK-2
    • dephosphorylation by phosphoprotein phosphatase (in response to insulin) activates PRK-2, and inactivates FBPase-2
    • Xylulose 5-phosphate (pentose pathway) allosterically upregulates phosphoprotein phosphatase
  • PFK-2/FBPase-2 isoenzyme response
    • key enzymes are differentially regulated in these tissues
    • Liver isozyme = phosphorylation on Ser 32 activates FBPase-2
  • Cardiac muscle isozyme = phosphorylation on Ser 406 and Thr 475 activates PFK-2

Steps 1 and 10

  • reciprocally regulated by acetyl-CoA
  • PK allosterically activated by F1,6-BP = first molecules committed to glycolysis
  • Pyruvate kinase = allosterically inhibited by:
    • ATP
    • Acetyl-CoA
    • Long chain fatty acids
    • Alanine
  • Liver has different pyruvate kinase isoform
    • phosphorylated by PKA in response to the hormone glucagon (which signals low blood sugar)
    • Slows liver PK, reserving scarce sugar for organs that need it
  • Pyruvate carboxylase (acetyl-CoA (+))
  • PEP carboxykinase (ADP(-))

5. Pentose Phosphate Pathway

1. Oxidative

  • Oxidation of glucose 6-P
  • produces NADPH from reductive biosynthesis
  • produces ribose 5-phosphate for the synthesis of nucleotides

2. Non-Oxidative

  • Glycolic intermediates
  • glucose 6-phosphate
    • Glucose → Glucose 6-phosphate dehydrogenase
    • stimulated by NADP+ (homotropic )
    • inhibited by NADPH (heterotropic)
    • Regulated by redox state of cytosol
  • xylulose 5-phosphate (modulator of phosphatase that stimulates liver PFK-2)
  • Transaldolase/Transketolase transfer 2 or 3 carbon atoms between sugar phosphates = rearrange 5C molecules into 6C moleules
  • 3C sugars are glyceraldehyde=3P → turned into Glucose-6P by gluconeogenesis
    • 6 X 5C → 5 X 6C

6. Glycogen Metabolism

  • Glycogen: polymer of glucose (storage)
    • Quick source of energy
  • Liver: glycolysis + control of blood glucosse
  • Muscle: glycolysis

1. Synthesis

  • Making the precursor

    1. glucose 6-P → glucose 1-P (phosphoglucomutase)
    2. glucose 1-P + UTP → UDP-glucose + PPi (UDP-glucose pyrophophorylase)
    • UDP-glucose then acts as an activated sugar donor

2. Glycogenolysis

  • Glycogen Phosphorylase
    • Glycogen -→ glucose + Glucose 1-phosphate (glycogen phosphorylase)
    • Glucose binding of phosporylase = allosteric = dephosphorylation by phosphorylase a phosphatase

3. Regulation of glycogen synthase and phosphorylase

  • Synthase = inhibited by phosphorylation (PKA and GSK3)

  • Phosphorylase = activated by phsphorylation

    • Phosphorylase B kinase = phosphorylates glycogen phosphorylase (activates it)

    • Glycogen phosphorylase a phosphatase (dephosphorylates)

    • REGULATED BY = PKA-mediated phosphorylation

7. Regulation by PKA and hormones

  • PKA regulation = low blood sugar leads to increased glycogen breakdown and decreased glycogen synthesis and glycolysis

  • Hormone-regulated enzyme activity coordinates tissue specific metabolism

    • Why are muscles and liver metabolism different?
    • Muscle lacks glucagon receptors
    • muscle pyruvate kinase = not phosphorylated by PKA
    • muscle lacks gluconeogenic enzymes
    • muscle lacks a key enzyme for glucose export
      • uses stored glycogen and glucose for itself