Cofactors & Vitamins
Andy Howard
Introductory Biochemistry
11 November 2008
Biochem: Cofactors,Vitamins 11/11/2008
What we’ll discuss
Coenzymes Vitamins
NAD and NADP Why they exist
FAD and FMN Water-soluble
Coenzyme A vitamins
TPP, PLP Cofactor precursors
Biotin, THF Ascorbate
Cobalamin Fat-Soluble
Lipoamide Vitamins
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Major cosubstrates (review)
Facilitate group transfers, mostly small groups
Oxidation-reduction participants
Cosubstrate Source Function
ATP Transfer P,Nucleotide
S-adenosylMet Methyl transfer
UDP-glucose Glycosyl transfer
NAD,NADP Niacin 2-electron redox
Coenzyme A Pantothenate Acyl transfer
Tetrahydrofolate Folate 1Carbon transfer
Ubiquinone Lipid-soluble e- carrier
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Major prosthetic groups (rvw)
Transfer of larger groups
One- or two-electron redox changes
Prosth.gp. Source Function
FMN, FAD Riboflavin 1e- and 2e- redox transfers
TPP Thiamine 2-Carbon transfers with C=O
PLP Pyridoxine Amino acid group transfers
Biotin Biotin Carboxylation, COO- transfer
Adenosyl- Cobalamin Intramolec. rearrangements
cobalamin
MeCobal. Cobalamin Methyl-group transfers
Lipoamide Transfer from TPP
Retinal Vitamin A Vision
Vitamin K Vitamin K Carboxylation of glu residues
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NAD+ and NADP+
Net charge isn’t really >0 ;
the + is just a reminder that the
nicotinamide ring is positively charged
Most important cosubstrates in oxidation-
reduction reactions in aerobic organisms
Structure courtesy of
Sergio Marchesini, U.
Brescia
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Differences between them
The chemical difference is in the
phosphorylation of the 2’ phosphate
group of the ribose moiety
The functional difference is that NAD+
is usually associated with catabolic
reactions and NADP+ is usually
associated with anabolic reactions
Therefore often NAD+ and NADPH are
reactants and NADH and NADP+ are
products
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How do we get back to the
starting point?
NADH is often oxidized back to NAD+ as
part of the electron-transport chain
Imbalances can be addressed via:
NAD Kinase (S.Kawai et al (2005),
J.Biol.Chem. 280:39200)
NADP phosphatase
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iClicker quiz: question 1
What would you expect to be the
phosphate donor in the NAD kinase
reaction?
(a) free phosphate
(b) pyrophosphate
(c) ATP
(d) pyridoxal phosphate
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Reduced forms of NAD(P)
Reduction occurs on the
nicotinamide ring
Ring is no longer net-
positive
Ring is still planar but
the two hydrogens on
the para carbon are not
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FAD and FMN
Flavin group based on riboflavin
Alternate participants in redox reactions
Prosthetic groups: tightly but noncovalently
bound to their enzymes
That protects against wasteful reoxidation of
reduced forms
FADH2 is weaker reducing agent than NADH
These are capable of one-electron oxidations
and reductions
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FAD and FMN structures
FAD has an AMP attached P to P
Structure courtesy
Paisley University
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FMN/FAD redox forms
Two-electron version: H+ + :H- transferred
Reaction diagram courtesy of Eric
Neeno-Eckwall, Hamline University
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(ADP-3’P)
Coenzyme A
Reactive portion
is free sulfhydryl
at one end of
the molecule (Pantoate) 2-mercapto-
Can form ethylamine)
thioester with
acetate, etc. b-alanine)
Pantoate + Structure courtesy of
b-alanine = MPB project, George
pantothenate Washington University
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Thiamine Pyrophosphate
Based on thiamine, vitamin B1
Many carboxylases and oxidative
decarboxylases use this coenzyme
So do transketolases (move 2 carbons
at a time between sugars with keto
groups)
Thiazolium ring is reactive center:
pKa drops from 15 in H2O to 6 in
enzyme
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TPP reactions
pyrimidine
thiazolium
Diagram courtesy of
Oklahoma State U.
Biochemistry program
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Pyridoxal
phosphate
PLP is prosthetic group for many
amino-acid-related enzymes,
particularly transaminations
Carbonyl group of PLP bound as a
Schiff base (imine) to -amino
group of lysine at active site
First step is always formation of
external aldimine; goes through
gem-diamine intermediate to
internal aldimine
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Transaminations
-amino acid1 +
-ketoacid2
-ketoacid1 +
-amino acid2
Example:
-amino acid1 = glutamate
-ketoacid2 = pyruvate
-ketoacid1 = a-ketoglutarate
-amino acid2 = alanine
Many biosynthetic / degradative pathways for
normal amino acids depend on these reactions
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Biotin
Rarity: vitamin is the prosthetic group
Used in reactions that transfer carboxyl
groups
… and in ATP-dependent carboxylations
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Biotin reactivity
Covalently bound to active-site lysines to
form species called biocytin
Pyruvate carboxylase is characteristic
reaction:
Diagram courtesy
University of Virginia Biochemistry
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Tetrahydrofolate
Primary donor of one-carbon units
(formyl, methylene, methyl)
Supplies methyl group for thymidylate
Dihydrofolate reductase (DHFR) is an
interesting drug target
Methotrexate as cancer chemotherapeutic:
cancer needs more thymidylate than healthy cells
Trimethoprim as antibacterial:
Bacterial DHFR is somewhat different from
eucaryotic DHFR because bacteria derive DHF
from other sources; humans get it from folate
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THF structure and function
Figure courtesy
horticulture program,
Purdue
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Cobalamin
Largest B vitamin
Corrin ring structure related to heme but
missing one carbon in ring structure
Cobalt bound in core of ring system
Involved in enzymatic rearrangements
Catabolism of odd-chain fatty acids
Methylation of homocysteine
Reductive dehalogenation
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Adenosyl-
Cobalamin
Reactive
“Missing” carbon Co-C bond
Diagram courtesy of
Swiss Food News
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Lipoamide
Protein-bound form of lipoic acid
Contains five-membered disulfide ring
Covalently bound via amide to protein
lysine sidechain
Involved in swinging arm between active
sites in multienzyme complexes
Disulfides break periodically
Example: pyruvate dehydrogenase
complex
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Lipoamide 2e- reduction
Cf. Scheme 7.6: thioester starting point
Fig. Courtesy Biochem
and Biophysics
program, Rensselaer
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iClicker revisited
Which coenzyme would you expect
would be required for the reaction
oxaloacetate + glutamate
aspartate + -ketoglutarate?
(a) ascorbate
(b) PLP
( c) thiamine pyrophosphate
(d) NAD
(e) none of the above
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Many cofactors are
derived from vitamins
We justify lumping these two topics
together because many cofactors are
vitamins or are metabolites of vitamins.
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Vitamins: necessary
micronutrients that cannot be
synthesized internally
What’s a vitamin for one organism is not
for another
Primates and some rodents are the only
vertebrates that don’t synthesize
ascorbate
E.coli can make almost everything given
energy and sources of atoms
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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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Vitamins: broad classifications
Water-soluble vitamins
Coenzymes or coenzyme precursors
Non-coenzymic metabolites
Fat-soluble vitamins
Antioxidants
Other lipidic vitamins
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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
arachidonate aren’t considered vitamins
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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
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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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QuickTime™ and a
TIFF (Uncompressed) decompressor
are needed to see this picture.
Deficiency of niacin
Pellagra: dermatitis, diarrhea,
dementia
Still found in some diets that are low Image courtesy
history.nih.gov
in vitamin content
Humans can actually synthesize
nicotinamide from tryptophan; but
that’s often in short supply too
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QuickTime™ and a
TIFF (Uncompressed) decompressor
are needed to see this picture.
Deficiency of thiamine
Beriberi: primary symptoms
are in nervous system and Image courtesy
musculature answers.com
Polished rice is missing
thiamine;
rice hulls are rich in it
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Deficiency of cobalamin
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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Ascorbate
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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PTM role of ascorbate
(fig. 6.17)
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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Dietary deficiency QuickTime™ and a
TIFF (Uncompressed) decompressor
are needed to see this picture.
of ascorbate
Primary sources of
ascorbate are fruits,
particularly citrus, and Image courtesy
U.Cincinnati
green vegetables Medical School
Ascorbate deficiency’s
first symptom involves
collagen degradation,
leading to scurvy
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Scurvy in history
QuickTime™ and a
Shortage of green vegetables in TIFF (Uncompressed) decompressor
are needed to see this picture.
sailors’ diets meant scurvy was
rampant on shipboard until the
18th century
Success of English navy over
French 1760-1800 was partly due
to the introduction of limes in
English sailors’ diets 50 years
before the French caught on
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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
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Vitamin A (retinol)
3 forms varying in terminal polar group
Involved in signaling and receptors
b-carotene is nonpolar dimer
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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
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Vitamin D
Several related forms
Hormones involved in
Ca2+ regulation
Figure courtesy (cholecalciferol)
Cyberlipid
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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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