1. Andy Howard Introductory Biochemistry 19 November 2013
General Metabolism I
Andy Howard Introductory Biochemistry
19 November 2013
Biochemistry: Metabolism I
11/19/2013

2. Metabolism: the core of biochemistry
All of biology 402 will concern itself with the specific pathways of metabolism
Our purpose here is to arm you with the necessary weaponry
We’ll then cover nutritional biochemistry, which fits naturally into these topics because many vitamins are precursors of coenzymes.
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Biochemistry: Metabolism I

3. Biochemistry: Metabolism I
What we’ll discuss
Nutritional biochemistry
Macronutrients
Vitamins
Water-soluble
Lipidic
Vitamins as coenzyme precursors
Metabolism
Definitions
Pathways
Control
Feedback
Flux
Phosphorylation
Other PTMs
Evolution
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Biochemistry: Metabolism I

4. Biochemistry: Metabolism I
Almost ready to start the specifics (chapter 18)
Define it! Metabolism is the network of chemical reactions that occur in biological systems, including the ways in which they are controlled.
So it covers most of what we do here!
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Biochemistry: Metabolism I

5. Intermediary Metabolism
Metabolism involving small molecules
Describing it this way is a matter of perspective: Do the small molecules exist to give the proteins something to do, or do the proteins exist to get the metabolites interconverted?
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Biochemistry: Metabolism I

6. How similar are pathways in various organisms?
Enormous degree of similarity in the general metabolic approaches all the way from E.coli to elephants
Glycolysis arose prior to oxygenation of the atmosphere
This is considered strong evidence that all living organisms are derived from a common ancestor
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Biochemistry: Metabolism I

7. Anabolism and catabolism
Anabolism: synthesis of complex molecules from simpler ones
Generally energy-requiring
Involved in making small molecules and macromolecules
Catabolism: degradation of large molecules into simpler ones
Generally energy-yielding
All the sources had to come from somewhere
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Biochemistry: Metabolism I

8. Common metabolic themes
Maintenance of internal concentrations of ions, metabolites, & (? enzymes)
Extraction of energy from external sources
Pathways specified genetically
Organisms & cells interact with their environment
Constant degradation & synthesis of metabolites and macromolecules to produce steady state
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Biochemistry: Metabolism I

9. Biochemistry: Metabolism I
Metabolism and energy
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Biochemistry: Metabolism I

10. Metabolic classifications
Carbon sources
Autotrophs vs. heterotrophs
Atmospheric CO2 as a C source vs. otherwise-derived C sources
Energy sources
Phototrophs vs. chemotrophs
(Sun)light as source of energy vs. reduced organic compounds as a source of energy
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Biochemistry: Metabolism I

11. Fourway divisions (table 17.2)
Energy/Carbon
Phototrophs: Energy from light
Chemotrophs: Energy from reduced organic molecules
Autotrophs: Carbon from atmospheric CO2
Photoautotrophs: Green plants, cyanobacteria, …
Chemoautotrophs: Nitrifying bacteria, H, S, Fe bacteria
Heterotrophs: Carbon from other [organic] sources
Photoheterotrophs: Nonsulfur purple bacteria
Chemoheterotrophs: Animals, many microorganisms, . . .
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Biochemistry: Metabolism I

12. Another distinction: the organism’s interaction with oxygen
Aerobes: use O2 as the ultimate electron acceptor in oxidation-reduction reactions
Anaerobes: don’t depend on O2
Obligate: poisoned by O2
Facultative: can switch hit
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Biochemistry: Metabolism I

13. Biochemistry: Metabolism I
Flow of energy
Sun is ultimate source of energy
Photoautotrophs drive synthesis of [reduced] organic compounds from atmospheric CO2 and water
Chemoheterotrophs use those compounds as energy sources & carbon; CO2 returned to atmosphere
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Biochemistry: Metabolism I

14. How to anabolism & catabolism interact?
Sometimes anabolism & catabolism occur simultaneously.
How do cells avoid futile cycling?
Just-in-time metabolism
Compartmentalization:
Anabolism often cytosolic
Catabolism often mitochondrial
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Biochemistry: Metabolism I

15. Biochemistry: Metabolism I
Pathway
A sequence of reactions such that the product of one is the substrate for the next
Similar to an organic synthesis scheme (but with better yields!)
May be:
Unbranched
Branched
Circular
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Biochemistry: Metabolism I

16. Biochemistry: Metabolism I
Catabolism stages
Stage 1: big nutrient macromolecules hydrolyzed into their building blocks
Stage 2: Building blocks degraded into limited set of simpler intermediates, notably acetyl CoA
Stage 3: Simple intermediates are fed to TCA cycle and oxidative phosphorylation
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Biochemistry: Metabolism I

17. Biochemistry: Metabolism I
Anabolism stages
Short list of simple precursors
These are elaborated in characteristic ways to build monomers e.g.: transamination of -ketoacids to make -amino acids
Those are then polymerized to form proteins, polysaccharides, polynucleotides, etc.
transamination
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18. Some intermediates play two roles
Some metabolites play roles in both kinds of pathways
We describe them as amphibolic
Just recall that: catabolism is many down to few, anabolism is few up to many
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Biochemistry: Metabolism I

19. Anapleurotic reactions
Certain metabolites that are integral to particular pathways also get used in other places
The result is that the concentration of those metabolites may get depleted by the competing pathways
The cell may need to replenish those metabolites in order to keep the original pathway humming
We describe reactions that replenish metabolites as anapleurotic
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Biochemistry: Metabolism I

20. Differences between catabolic and anabolic pathways
Often they share many reactions, notably the ones that are nearly isoergic (Go ~ 0)
Reactions with Go < -20 kJ mol-1 are not reversible as is
Those must be replaced by (de)coupled reactions so that the oppositely-signed reactions aren’t unfeasible
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Biochemistry: Metabolism I

21. Other differences involve regulation
Generally control mechanisms influence catalysis in both directions
Therefore a controlling influence (e.g. an allosteric effector) will up- or down-regulate both directions
If that’s not what the cell needs, it will need asymmetric pathways or pathways involving different enzymes in the two directions
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Biochemistry: Metabolism I

22. Biochemistry: Metabolism I
ATP’s role
We’ve discussed its significance as an energy currency
It’s one of two energy-rich products of the conversion of light energy into chemical energy in phototrophs
ATP then provides drivers for almost everything else other than redox
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Biochemistry: Metabolism I

23. Biochemistry: Metabolism I
NAD’s role
NAD acts as as an electron acceptor via net transfer of hydride ions, H:-, in catabolic reactions
Reduced substrates get oxidized in the process, and their reducing power ends up in NADH
Energy implied by that is used to make ATP (2.5 ATP/NAD) in oxidative phosphorylation
Image courtesy Michigan Tech Biological Sciences
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Biochemistry: Metabolism I

24. Biochemistry: Metabolism I
NADPH’s role
Involved in anabolic redox reactions
Reducing power in NADPH  NADP used to reduce some organic molecule
Involves hydride transfers again
NADPH regenerated in phototrophs via light-dependent reactions that pull electrons from water
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Biochemistry: Metabolism I

25. How to detect NAD reactions
340 nm
Absorbance
NADH
NAD+ and NADH (and NADP+ and NADPH) have extended aromatic systems
But the nicotinamide ring absorbs strongly at 340 only in the reduced (NADH, NADPH) forms
Spectrum is almost pH-independent, too!
So we can monitor NAD and NADP-dependent reactions by appearance or disappearance of absorption at 340 nm
Wavelength
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Biochemistry: Metabolism I

26. How do we study pathways?
Inhibitor studies
Mutagenesis
Isotopic tracers (radio- or not)
NMR
Disruption of cells to examine which reactions take place in which organelle
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Biochemistry: Metabolism I

27. Why multistep pathways?
Limited reaction specificity of enzymes
Control of energy input and output:
Break big inputs into ATP-sized inputs
Break energy output into pieces that can be readily used elsewhere
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Biochemistry: Metabolism I

28. iClicker quiz question 1
A reaction A+B  C+D proceeds from left to right in the cytosol and from right to left in the mitochondrion. As written, it is probably
(a) a catabolic reaction
(b) an anabolic reaction
(c) an amphibolic reaction
(d) we don’t have enough information to answer.
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Biochemistry: Metabolism I

29. iClicker quiz question 2
An asymmetry between stage 1 of catabolism (C1) and the final stage of anabolism (A3) is
(a) A3 always requires light energy; C1 doesn’t
(b) A3 never produces nucleotides; C1 can involve nucleotide breakdown
(c) A3 adds one building block at a time to the end of the growing polymer; C1 can involve hydrolysis in the middle of the polymer
(d) There are no asymmetries between A3 and C1
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Biochemistry: Metabolism I

30. iClicker quiz question 3
Could dAMP, derived from degradation of DNA, serve as a building block to make NADP?
(a) Yes.
(b) Probably not: the energetics wouldn’t allow it.
(c) Probably not: the missing 2’–OH would make it difficult to build NADP
(d) No: dAMP is never present in the cell
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Biochemistry: Metabolism I

31. Biochemistry: Metabolism I
Nutrition
Lots of nonsense, some sense on this subject
Skepticism among MDs as to its relevance
Fair view is that nutrition matters in many conditions, but it’s not the only determinant of health
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Biochemistry: Metabolism I

32. Biochemistry: Metabolism I
Macronutrients
Proteins
Carbohydrates
Lipids
Fiber
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33. Biochemistry: Metabolism I
Protein as food
Source of essential amino acids
Source of non-essential amino acids
Fuel (often via interconversion to -ketoacids and incorporation into TCA)
All of the essential amino acids must be supplied in adequate quantities
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Biochemistry: Metabolism I

34. Which amino acids are essential?
At one level, that’s an easy question to answer: they’re the ones for which we lack a biosynthetic pathway: KMTVLIFWH
That shifts the question to: why have some of those pathways survived and not all?
Answer: pathways that are complex or require more than ~30 ATP / aa are absent (except R,Y)
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Biochemistry: Metabolism I

35. Essential & non-essential aa’s
A.A. name # ATP’s
Glycine 12
Serine 18
Cysteine 19
Alanine 20
Aspartate 21
Asparagine 22-24
Glutamate 30
Glutamine 31
Proline 39
Arginine 44 *
Tyrosine 62 **
Essential
A.A. Name #ATPs
Threonine 31
Valine 39
Histidine 42
Methionine 44
Leucine 47
Lysine 50-51
Isoleucine 55
Phenylalanine 65
Tryptophan 78
* Essential in some organisms ** Derived from phenylalanine
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Biochemistry: Metabolism I

36. Biochemistry: Metabolism I
Carbohydrates as food
Generally recommended to be more than half of caloric intake
Complex carbohydrates are hydrolyzed to glucose-1-P and stored as glycogen or interconverted into other metabolites
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Biochemistry: Metabolism I

37. Biochemistry: Metabolism I
Lipids as food
You’ll see in 402 that the energy content of a lipid is ~ 2x that of carbohydrates simply because they’re more reduced
They’re also more efficient food storage entities than carbs because they don’t require as much water around them
Certain fatty acids are not synthesizable; by convention we don’t call those vitamins
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Biochemistry: Metabolism I

38. Biochemistry: Metabolism I
Vitamins
Vitamins are necessary micronutrients
A molecule that is a vitamin in one organism isn’t necessarily a vitamin in another
E.coli can make all necessary metabolites given sources of water, nitrogen, and carbon
Most eukaryotic chemoautotrophs find it more efficient to rely on diet to make complex metabolites
We’ll discuss water-soluble vitamins first, then lipid vitamins
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Biochemistry: Metabolism I

39. Why wouldn’t organisms make everything?
Complex metabolites require energy for synthesis
Control of their synthesis is also metabolically expensive
Cheaper in the long run to derive these nutrients from diet
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Biochemistry: Metabolism I

40. Vitamins: broad classifications
Water-soluble vitamins
Coenzymes or coenzyme precursors
Non-coenzymic metabolites
Fat-soluble vitamins
Antioxidants
Other lipidic vitamins
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41. Are all nutrients that we can’t synthesize considered vitamins?
No:
If it’s required in large quantities, it’s not a vitamin
By convention, essential fatty acids like linoleate aren’t considered vitamins
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Biochemistry: Metabolism I

42. Warning: ugly photos coming
I have included some web-derived photos of patients with severe vitamin deficiencies
If you’re squeamish, be prepared.
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Biochemistry: Metabolism I

43. Vitamins: broad classifications
Water-soluble vitamins
Coenzymes or coenzyme precursors
Non-coenzymic metabolites
Fat-soluble vitamins
Antioxidants
Other lipidic vitamins
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Biochemistry: Metabolism I

44. Are all nutrients that we can’t synthesize considered vitamins?
No:
If it’s required in large quantities, it’s not a vitamin
By convention, essential fatty acids like linoleate aren’t considered vitamins
11/19/2013
Biochemistry: Metabolism I

45. Warning: ugly photos coming
I have included some web-derived photos of patients with severe vitamin deficiencies
If you’re squeamish, be prepared.
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Biochemistry: Metabolism I

46. Biochemistry: Metabolism I
Coenzyme precursors
We’ve already outlined the fact that most water-soluble coenzymes are derived from vitamins—typically B vitamins
Typically the dietary form can be converted by a fairly short metabolic pathway into the coenzyme form, e.g.
niacin + glutamine  nicotinamide + glutamate
nicotinamide + ADP-ribose  NAD
Some coenzyme precursors are, in fact, lipidic
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Biochemistry: Metabolism I

47. Biochemistry: Metabolism I
The B vitamins
All aqueous micronutrients
Generally identified via pathologies associated with dietary deficiencies
B1: thiamin (produces TPP)
B2: riboflavin (produces FAD, FMN)
B3: niacin (produces NAD, NADP)
B5: pantothenate (produces Coenzyme A)
B6: pyridoxamine (produces PLP)
B9: folate: produces THF, THF derivatives
B12: cobalamin (produces adenosylcobalamin, methylcobalamin)
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Biochemistry: Metabolism I

48. Deficiency of niacin (B3)
Remember: niacin is the source for NAD and NADP (redox cofactors)
Pellagra: dermatitis, diarrhea, dementia
Still found in some diets that are low in vitamin content
Humans can actually synthesize nicotinamide from tryptophan; but that’s often in short supply too
Image courtesy history.nih.gov
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Biochemistry: Metabolism I

49. Deficiency of thiamin (B1)
Remember: thiamin is precursor to TPP (used in decarboxylations)
Beriberi: primary symptoms are in nervous system &musculature
Polished rice is missing thiamine; rice hulls are rich in it
Image courtesy answers.com
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Biochemistry: Metabolism I

50. Riboflavin (Vitamin B2)
Precursor of the redox cofactors flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD)
Key property: can undergo one-electron as well as two-electron redox interactions because of delocalization across the isoalloxazine aromatic ring system
Deficiencies can lead to growth retardation
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Biochemistry: Metabolism I

51. Pantothenate (Vitamin B5)
Precursor of coenzyme A, critical in lipid and TCA-cycle metabolism
Made up of pantoate and beta-alanine components
Deficiency leads to dermatitis in chickens
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Biochemistry: Metabolism I

52. Biochemistry: Metabolism I
pyridoxine
Pyridoxal (Vitamin B6)
Precursor of pyridoxal phosphate (PLP), a crucial cofactor in enzymatic reactions involving amino acids, including transaminations
Available in diet as pyridoxine, pyridoxal, pyridoxamine
Modest deficiencies lead to dermatitis in rats
pyridoxal
pyridoxamine
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Biochemistry: Metabolism I

53. Biochemistry: Metabolism I
Biotin (Vitamin B7)
Used directly as a coenyzme in carboxylation reactions
Modest deficiency leads to dermatitis in humans
Most hydrophobic of the B vitamins
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54. Biochemistry: Metabolism I
Folate (Vitamin B9)
Precursor to tetrahydrofolate, N5,N10-methylene THF, other active cofactor forms
Deficiency leads to anemia
Critical to creation of thymidylate, so it’s important to growth in fetus
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Biochemistry: Metabolism I

55. Deficiency of cobalamin (B12)
Available sources of cobalamin are animal products
Therefore vegans need to be careful to get cobalamin from supplements
Symptoms of deficiency (pernicious anemia): weakness, fatigue, pallor, palpitations, dizziness
Deficiency is common even in non-vegans: 5-40% of the population?
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Biochemistry: Metabolism I

56. Biochemistry: Metabolism I
How do herbivores get B12?
Strict herbivores like cattle do require cobalamin: how do they get it?
Answer: gut bacteria in the cattle produce enough for the cattle to use
This illustrates the fact that even large animals don’t need much of this
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Biochemistry: Metabolism I

57. Biochemistry: Metabolism I
iClicker question 4
4. Why have we not outlined a disease associated with deficiencies of vitamin B6 (pantothenate)?
(a) Vitamin B3 can be converted to B6.
(b) The disease is so rapid in onset that no one ever had a reason to name it.
(c) Almost any normal diet contains adequate quantities of B6.
(d) The name for the deficiency is unpronounceable.
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58. Biochemistry: Metabolism I
Ascorbate (C)
Vitamin in primates, some rodents
Synthesizable in most other vertebrates
Involved in collagen processing
Reduced form acts as reducing agent during hydroxylation of collagen
Deficiency gives rise to inadequate collagen - scurvy
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Biochemistry: Metabolism I

59. PTM role of ascorbate (G&G fig. 6.15)
Proline + O2 + -ketoglutarate + ascorbate  4-hydroxyproline + succinate + CO2 + dehydroascorbate
This is a post-translational modification that occurs to prolines within collagen
The hydroxylated prolines help stabilize the collagen triple helix
Hydroxylysine found in collagen too
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Biochemistry: Metabolism I

60. Dietary deficiency of ascorbate
Primary sources of ascorbate are fruits, particularly citrus, and green vegetables
Ascorbate deficiency’s first symptom involves collagen degradation, leading to scurvy
Image courtesy U. Cincinnati Medical School
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Biochemistry: Metabolism I

61. Biochemistry: Metabolism I
Scurvy in history
Shortage of green vegetables in sailors’ diets meant scurvy was rampant on shipboard until the 18th century
Success of English navy over French was partly due to the introduction of limes in English sailors’ diets 50 years before the French caught on
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Biochemistry: Metabolism I

62. Biochemistry: Metabolism I
Lipid vitamins
Contain rings & long aliphatic sidechains
At least one polar group in each
Absorbed in intestine, carried via bile salts
Hard to study
Most are formally built from isoprene units, as are steroids
2-methyl- 1,3-butadiene
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Biochemistry: Metabolism I

63. Biochemistry: Metabolism I
Vitamin A (retinol)
3 forms varying in terminal polar group; based on four isoprene units
Involved in signaling and receptors
b-carotene is nonpolar dimer
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Biochemistry: Metabolism I

64. Biochemistry: Metabolism I
Vitamin A deficiency
Produces night blindness because the retina and cornea dry out
Most common cause: nursing infants whose mothers have vitamin A deficiency in their diet
Corneal scarring due to vitamin A deficiency:
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Biochemistry: Metabolism I

65. Biochemistry: Metabolism I
Vitamin D
Vitamin D3 (cholecalciferol)
Several related forms
Cholesterol-derived C30 (6 isoprenes)
Hormones involved in Ca2+ regulation
Cancer chemoprevention!
Vitamin D2 (ergocalciferol)
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66. Biochemistry: Metabolism I
Vitamin D deficiency
Rickets in children: Bone disease, restlessness, slow growth
One form of vitamin D is actually synthesizable from cholesterol given adequate sunlight;
Therefore rickets is most common in densely settled urban environments
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Biochemistry: Metabolism I

67. Vitamin E (a-tocopherol)
C30 (6 isoprenes) nonsteroidal vitamin
Phenol can undergo 1e- oxidation to moderately stable free radical
Antioxidant activity prevents damage to fatty acids in membranes
α-tocopherol
phenol
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Biochemistry: Metabolism I

68. Vitamin K (phylloquinone)
C30 (6 isoprenes) non-steroidal vitamin
Involved in synthesis of proteins involved in blood coagulation
Reduced form involved as reducing agent in carboxylation reaction on glu sidechains
phylloquinone
phylloquinone 2,3-epoxide
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Biochemistry: Metabolism I