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Amino acid
catabolism;
Nucleic acid chemistry
Andy Howard
Introductory Biochemistry
29 April 2008
What we’ll cover

Amino acid catabolism  Nucleosides




Urea cycle



Degradation products  Nucleotides
Interconversions
 Oligo- and
Specifics
polynucleotides
Reactions
Cellular localization
Nucleic acid chemistry


Pyrimidines: C, U, T
Purines: A, G
Amino Acid metabolism



Duplex DNA
Helicity
RNA


structure
types
p. 2 of 65
29 April 2008
What do we do with
amino acids?



Obviously a lot of them serve as building-blocks
for protein and peptide synthesis via ribosomal
mechanisms
Also serve as metabolites, getting converted to
other compounds or getting oxidized as fuel
Most amino acid degradations begin with
transaminations to make glutamate; the resulting
alpha-keto acids are further metabolized
Amino Acid metabolism
p. 3 of 65
29 April 2008
Transaminations

Generally two stages:



amino acid + -ketoglutarate 
-keto acid + glutamate
Glutamate + NAD+ + H2O 
-ketoglutarate + NADH + H+ + NH4+
Net reaction is
amino acid + NAD+ + H2O 
-keto acid + NADH + H+ + NH4+
Amino Acid metabolism
p. 4 of 65
29 April 2008
Glucogenic and
ketogenic amino acids

Degradation of many amino acids lead to
TCA cycle intermediates or pyruvate



Degradation of others leads to acetyl CoA
and related compounds



therefore these can be built back up to glucose;
these are called glucogenic
these cannot be built back up to glucose except
via the glyoxalate shuttle
these are called ketogenic
Some amino acids are both!
Amino Acid metabolism
p. 5 of 65
29 April 2008
Glucogenic amino acids


Amino acids that can be catabolized to
produce building blocks that lead to
glucose without help of glyoxalate
pathway
Most produce succinate, succinyl CoA,
fumarate, a-ketoglutarate, or
oxaloacetate
Amino Acid metabolism
p. 6 of 65
29 April 2008
Ketogenic amino acids


These do not produce TCA cycle
intermediates, but rather produce acetyl
CoA or its close relatives
Can be built back up into fats or ketone
bodies
Amino Acid metabolism
p. 7 of 65
29 April 2008
Serine-based metabolites


Serine is a building block for sphinganine
and therefore for sphingolipids
Serine also leads to phosphatidylserine,
which is important by itself and can be
metabolized to phosphatidylethanolamine
and phosphatidylcholine
Amino Acid metabolism
p. 8 of 65
29 April 2008
Serine
degradation

Two paths for degrading serine:


PLP-dependent serine dehydratase
simply deaminates ser to pyruvate;
this enzyme is like trp synthase
More common: SHMT transfers
hydroxymethyl group to THF, leaving
glycine; we’ve seen that one as a
biosynthetic enzyme for making glycine
Amino Acid metabolism
p. 9 of 65
Serine
dehydratase
PDB 1P5J
41 kDa
monomer
human
29 April 2008
Glycine-based
metabolites

porphobilinogen
Glycine is a source for purines,
glyoxalate, creatine phosphate, and (with
the help of succinyl CoA)
porphobilinogen, whence we get
porphyrins, and from those we get
chlorophyll, heme, and cobalamin
Amino Acid metabolism
p. 10 of 65
29 April 2008
Glycine cleavage
system



Glycine + H2O + NAD+ + THF 
NADH + H+ + HCO3- + NH4+ +
5-10-methyleneTHF
Complex system: PLP,
lipoamide, FAD prosthetic groups
Lipoamide swinging arm works
as in pyruvate dehydrogenase
Amino Acid metabolism
p. 11 of 65
T protein
(aminomethyltransferase)
of glycine
cleavage
system
PDB 1V5V
88 kDa dimer
Pyrococcus
29 April 2008
asp, glu, ala degradation

Standard transmination converts aspartate to
oxaloacetate with release of glutamate, which
then can be deaminated to re-form ketoglutarate:




asp + -kg  oxaloacetate + glu
glu + NAD+ + H2O  -kg + NADH + H+ + NH4+
Deamination converts glutamate to ketoglutarate, as above
Standard transamination converts alanine to
pyruvate according to the same logic as asp
Amino Acid metabolism
p. 12 of 65
29 April 2008
All three of these are
glucogenic!


-ketoglutarate and oxaloacetate are
TCA cycle intermediates
Pyruvate feeds the TCA cycle in ways
that can lead to glucose
Amino Acid metabolism
p. 13 of 65
29 April 2008
Degradation of
asn, gln




Asparagine and glutamine are
deaminated to asp and glu
Thus they lead to oxaloacetate and
-ketoglutarate, respectively
So they’re glucogenic
The initial hydrolyses (deaminations)
are catalyzed by asparaginase and
glutaminase
Amino Acid metabolism
p. 14 of 65
Asparaginase
PDB 1O7J
144 kDa
tetramer
Erwinia
chrysanthemi
29 April 2008
Arginine
degradation



Arginine is hydrolyzed to urea
and ornithine as part of the
urea cycle; enzyme is arginase
PLP-dependent enzyme
converts ornithine to glu semialdehyde
That’s oxidized to glutamate
Amino Acid metabolism
p. 15 of 65
Arginase
PDB 2AEB
212 kDa hexamer
Dimer shown
Human
29 April 2008
Proline
degradation

Proline oxidized back to
Pyrroline 5-carboxylate



 1-
O2 is oxidizing agent
different enzyme from forward
reaction
Proline
dehydrogenase
PDB 2EKG
72 kDa dimer
Thermus
thermophilus
Ring opened non-enzymatically
to form glutamate semialdehyde; see arginine
Amino Acid metabolism
p. 16 of 65
29 April 2008
urocanate
Histidine
degradation



3 reactions from histidine to
N-formiminoglutamate;
first (HAL) makes urocanate
from histidine
Tetrahydrofolate-dependent reaction
produces glutamate and 5formiminoTHF
5-formiminoTHF is enzymatically
deaminated to 5,10-methyleneTHF,
which can be used in purine
synthesis, etc.
Amino Acid metabolism
p. 17 of 65
Histidine-ammonia
lyase
PDB 1GKM
224 kDa tetramer
monomer shown
Pseudomonas
putida
29 April 2008
How are we doing so far?



We did ser and gly first because they’re
so important
Then we’ve done a whole bunch that
connect up to glutamate (or asp):
asp, glu, ala, asn, gln, arg, pro, his
So we’re halfway through.
Amino Acid metabolism
p. 18 of 65
29 April 2008
Threonine
degradation





Several pathways (fig. 17.29)
Major one: oxidize threonine to
2-amino-3-ketobutyrate
2-amino-3-keto-butyrate reacts
with HS-CoA to form acetyl CoA
and glycine
So this one is ketogenic
Other pathways are glucogenic
Amino Acid metabolism
p. 19 of 65
Threonine
dehydrogenase
PDB 2DFV
115 kDa trimer
Pyrococcus
29 April 2008
Valine degradation
(fig. 17.30, center)


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
Valine transaminated to
-ketoisovalerate
Branched-chain -keto acid dehydrogenase
(TTP, Lipoamide):
-ketoisoavalerate + NAD+ + HS-CoA 
a-ketoisovaleryl CoA
Next reaction (acyl CoA dehydrogenase) 
2-methyl-1-propenyl CoA + NADH + CO2
Product undergoes 4 reactions to propionyl
CoA and thence to succinyl CoA: glucogenic
Amino Acid metabolism
p. 20 of 65
PDB 2VBF
125 kDa
dimer
Lactococcus
29 April 2008
Isoleucine and leucine
degradation



Same path but products are:
Leucine’s products: acetyl CoA +
acetoacetate: ketogenic
Isoleucine: Acetyl CoA + propionyl CoA:
ketogenic and glucogenic
Amino Acid metabolism
p. 21 of 65
29 April 2008
Methionine
degradation

A lot of methionine is turned into
S-adenosylmethionine:

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
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Methyl donor
Leaves behind S-Adenosylhomocysteine
S-adenosylhomocysteine can be
hydrolyzed to homocysteine and water
Homocysteine can condense with serine
to form cystathionine, which can yield
cysteine and -ketobutyrate… and we
know how to turn -ketobutyrate into
propionyl CoA. So met is glucogenic.
Amino Acid metabolism
p. 22 of 65
29 April 2008
Cysteine
degradation

Cysteinesulfinate

Most common: oxidation to
cysteinesulfinate, which
transaminates to form sulfinylpyruvate:
cysteine + O2 
cysteinesulfinate + H+
-sulfinylpyruvate undergoes
nonenzymatic desulfuration
to SO2 and pyruvate. So
cysteine is glucogenic.
Amino Acid metabolism
p. 23 of 65
Cysteine
dioxygenase
PDB 2B5H
22 kDa
monomer
rat
29 April 2008
Tetrahydrobiopterin
Phenylalanine


Simple: phenylalanine
gets hydroxylated to
form tyrosine:
phenylalanine + O2 
tyrosine
This is a
tetrahydrobiopterindependent enzyme—a
folate-like cofactor
Amino Acid metabolism
Phenylalanine hydroxylase
PDB 1J8U
71 kDa dimer
monomer shown
human
(residues 103-427)
p. 24 of 65
29 April 2008
Phenylketonuria


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Usually associated with mutation in
phenylalanine hydroxylase:
Accumulated Phe  phenylpyruvate
Afflicts 1/15000 newborns
Built-up phenylpyruvate causes irreversible
mental retardation
Type IV PKU related to deficiencies in enzymes
that restore tetrahydrobiopterin (see fig. 17.33,
bottom)
Amino Acid metabolism
p. 25 of 65
29 April 2008
homogentisate
Tyrosine
degradation



Transaminated and
mutated to
homogentisate
Three more reactions
convert that to fumarate
+ acetoacetate
So tyr (and phe) are
both ketogenic and
glucogenic
Amino Acid metabolism
Homogentisate
dioxygenase
PDB 1EYB
311 kDa
hexamer
Monomer shown
Human
p. 26 of 65
29 April 2008
Tryptophan
degradation



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
Tryptophan: need to open 2 rings!
8 reactions lead to alanine and ketoadipate; first is
trp + O2 -> N-formyl-kynurenine
Alanine gets transaminated to
pyruvate
-ketoadipate goes through 6
more reactions to acetyl CoA +
2CO2
So it’s ketogenic and glucogenic
Amino Acid metabolism
p. 27 of 65
Indoleamine 2,3dioxygenase
PDB 2D0T
89 kDa dimer
monomer shown
human
29 April 2008
Lysine
degradation
(fig. 17.35)





Condense lysine with -ketoglutarate
to form saccharopine
That’s deglutamated (?), oxidized, and
transaminated to -ketoadipate
Six reactions degrade that to 2 acetyl
CoA molecules plus 2 CO2
Purely ketogenic
Some bacteria decarboxylate it to
cadaverine
Amino Acid metabolism
p. 28 of 65
Saccharopine
dehydrogenase
PDB 2AXQ
103 kDa dimer
monomer shown
Yeast
29 April 2008
The urea cycle:
overview



This is a significant pathway in the
eukaryotic management of nitrogencontaining compounds
It was also one of the first
biochemical pathways to be carefully
characterized—by Krebs and
coworkers!
Proceeds via ornithine & citrulline to
urea and (in some organisms) uric
acid
Amino Acid metabolism
p. 29 of 65
ornithine
urea
29 April 2008
Making carbamoyl
phosphate
(fig. 17.37)



Bicarbonate is phosphorylated to
form
Ammonia condenses with that to
form carbamate and Pi
Second ATP-phosphorylation
forms carbamoyl phosphate
Amino Acid metabolism
p. 30 of 65
29 April 2008
Urea cycle itself


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In mitochondrion: carbamoyl phosphate
condenses with ornithine to form citrulline
Citrulline condenses with urea to form
arginosuccinate
Arginosuccinate is cleaved nonhydrolytically to
fumarate and arginine
Arginine yields urea and citrulline
Citrulline re-enters cycle
Amino Acid metabolism
p. 31 of 65
29 April 2008
iClicker quiz: question 1

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1. Glutamate + ammonia  glutamine + H2O
is only slightly endergonic (Go’ = +14 kJ mol-1),
yet it is coupled to ATP hydrolysis. Why?
(a) You can never run a reaction with a positive
Go’
(b) [glutamate] ~ [glutamine] in the cell
(c) If you heat the substrates, they disintegrate
(d) ammonia is toxic in the absence of ATP
Amino Acid metabolism
p. 32 of 65
29 April 2008
iClicker quiz #2

2. Which ribosomal amino acid’s
biosynthesis is closely associated with
the urea cycle?





(a) alanine
(b) serine
(c) ornithine
(d) arginine
(e) none of the above.
Amino Acid metabolism
p. 33 of 65
29 April 2008
6
5
Pyrimidines






N
4
2
N
3
Single-ring nucleic acid bases
pyrimidine
6-atom ring; always two nitrogens in the ring,
meta to one another
Based on pyrimidine, although pyrimidine itself
is not a biologically important molecule
Variations depend on oxygens and nitrogens
attached to ring carbons
Tautomerization possible
Note line of symmetry in pyrimidine structure
Amino Acid metabolism
p. 34 of 65
1
29 April 2008
H
N
O
Uracil and thymine




Uracil is a simple dioxo
derivative of pyrimidine:
2,4-dioxopyrimidine
Thymine is 5-methyluracil
Uracil is found in RNA;
Thymine is found in DNA
We can draw other
tautomers where we move
the protons to the oxygens
Amino Acid metabolism
p. 35 of 65
O
HN
uracil
HN
O
N
H
thymine
29 April 2008
O
H
N
O
Tautomers


Lactam and
Lactim forms
Getting these right
was essential to
Watson & Crick’s
development of
the DNA double
helical model
Amino Acid metabolism
HN
O
NH
O
uracil - lactam
H
N
O
uracil - lactim
HN
HN
O
N
H
thymine - lactam
O
O
N
thymine - lactim
p. 36 of 65
29 April 2008
OH
H
N
O
Cytosine
NH2
N
cytosine




This is 2-oxo,4-aminopyrimidine
It’s the other pyrimidine base found in
DNA & RNA
Spontaneous deamination (CU)
Again, other tautomers can be drawn
Amino Acid metabolism
p. 37 of 65
29 April 2008
Cytosine:
amino and imino forms

Again, this tautomerization needs to be
kept in mind
H
N
O
NH2
N
cytosine -amino form
Amino Acid metabolism
H
N
O
NH
N
cytosine -imino form
p. 38 of 65
29 April 2008
7
6
5
1N
N
3



8
4
2
Purines
H
N
N
9
Derivatives of purine; again, the purine
root molecule isn’t biologically
important
Six-membered ring looks a lot like
pyrimidine
Numbering works somewhat
differently: note that the glycosidic
bonds will be to N9, whereas it’s to
N1 in pyrimidines
Amino Acid metabolism
p. 39 of 65
29 April 2008
Adenine




This is 6-aminopurine
Found in RNA and DNA
We’ve seen how important adenosine
and its derivatives are in metabolism
Tautomerization happens here too
NH
NH2
H
N
N
N
N
adenine - amino form
Amino Acid metabolism
H
N
HN
N
N
adenine - imino form
p. 40 of 65
29 April 2008
Guanine
This is 2-amino-6-oxopurine
 Found in RNA, DNA
 Lactam, lactim forms
OH

O
H
N
H
N
N
HN
H2N
N
guanine - lactam
Amino Acid metabolism
N
H2N
N
N
guanine - lactim
p. 41 of 65
29 April 2008
O
HO
Nucleosides
NR1R2
OH
HO
N-glycoside of ribofuranose



As mentioned in ch. 8, these are
glycosides of the nucleic acid bases
Sugar is always ribose or deoxyribose
Connected nitrogen is:


N1 for pyrimidines (on 6-membered ring)
N9 for purines (on 5-membered ring)
Amino Acid metabolism
p. 42 of 65
29 April 2008
Pyrimidine nucleosides

Drawn here in amino and lactam forms
OH
OH
HO
HO
OH
O
N
H2N
N
OH
O
O
N
H
cytidine
Amino Acid metabolism
O
N
O
uridine
p. 43 of 65
29 April 2008
Pyrimidine
deoxynucleosides
OH
OH
H
H
OH
O
N
O
N
H
OH
O
OH
2'-deoxyuridine
O
N
H
OH
N
O
2'-deoxythymidine
O
N
H2N
O
N
O
deoxycytidine
Amino Acid metabolism
p. 44 of 65
29 April 2008
A tricky nomenclature issue


Remember that thymidine and its
phosphorylated derivatives ordinarily
occur associated with deoxyribose, not
ribose
Therefore many people leave off the
deoxy- prefix in names of thymidine and
its derivatives: it’s usually assumed.
Amino Acid metabolism
p. 45 of 65
29 April 2008
Purine nucleosides

Drawn in amino and lactam forms
NH2
O
N
N
N
HN
N
N
H2N
N
N
O
O
HO
HO
OH
OH
HO
HO
guanosine
adenosine
Amino Acid metabolism
p. 46 of 65
29 April 2008
Purine deoxynucleosides
O
NH2
N
N
HN
N
N
H2N
N
N
N
O
O
OH
OH
HO
HO
deoxyguanosine
deoxyadenosine
Amino Acid metabolism
p. 47 of 65
29 April 2008
Chirality in nucleic acids





Bases themselves are achiral
Four asymmetric centers in
ribofuranose, counting the glycosidic
bond.
Three in deoxyribofuranose
Glycosidic bond is one of those 4 or 3.
Same for nucleotides:
phosphates don’t add asymmetries
Amino Acid metabolism
p. 48 of 65
29 April 2008
NH2
Monophosphorylated
nucleosides


N
N
We have specialized names for
the 5’-phospho derivatives of the
nucleosides, i.e. the nucleoside
monophosphates:
They are nucleotides




N
N
O
HO
OO
P
HO
O
adenylate
Adenosine 5’-monophosphate =
AMP = adenylate
GMP = guanylate
CMP = cytidylate
UMP = radiate
Amino Acid metabolism
p. 49 of 65
O-
29 April 2008
Deoxynucleotides

O
Similar nomenclature




dAMP =
deoxyadenylate
dGMP =
deoxyguanylate
dCMP =
deoxycytidylate
dTTP (= TTP) =
deoxythymidylate =
thymidylate
Amino Acid metabolism
N
HN
H2N
N
N
O
OO
OP
HO
O
deoxyguanylate
p. 50 of 65
29 April 2008
Di and triphosphates

Phosphoanhydride bonds link second and
perhaps third phosphates to the 5’-OH on
the ribose moiety
O
N
O
H2N
O
O
O
P
P
P
O
N
O
O-
O
O-
OH
O-
Mg2+
OH
HO
cytidine triphosphate
Amino Acid metabolism
p. 51 of 65
29 April 2008
Oligomers and Polymers



Monomers are nucleotides or
deoxynucleotides
Linkages are phosphodiester linkages
between 3’ of one ribose and 5’ of the next
ribose
It’s logical to start from the 5’ end for
synthetic reasons
Amino Acid metabolism
p. 52 of 65
29 April 2008
Typical DNA dinucleotide


Various notations: this is pdApdCp
Leave out the p’s if there’s a lot of them!
-O
OP
O
O
O
-O
N
O-
N
P
O
O
O
O
N
-O
P
O
HN
O
NH2
O
N
O
Amino Acid metabolism
p. 53 of 65
N
NH2
29 April 2008
DNA structure



Many years of careful
experimental work enabled
fabrication of double-helical
model of double-stranded
DNA
Explained [A]=[T], [C]=[G]
Specific H-bonds stabilize
double-helical structure:
see fig. 19.12
Amino Acid metabolism
p. 54 of 65
29 April 2008
What does double-stranded
DNA really look like?



Picture on previous slide emphasizes
only the H-bond interactions
Fig.19.12 is better: shows the tilt of the
sugars
Planes of the bases are almost
perpendicular to the helical axes on both
sides of the double helix
Amino Acid metabolism
p. 55 of 65
29 April 2008
Sizes (see fig. 19.14)




Diameter of the double helix: 2.37nm
Length along one full turn:
10.4 base pairs = pitch = 3.40nm
Distance between stacked base pairs =
rise = 0.33 nm
Major groove is wider and shallower;
minor groove is narrower and deeper
Amino Acid metabolism
p. 56 of 65
29 April 2008
What stabilizes this?

Variety of stabilizing
interactions




Stacking of base pairs
Hydrogen bonding between
base pairs
Hydrophobic effects (burying
bases, which are less polar)
Charge-charge interactions:
phosphates with Mg2+ and
cationic proteins
Amino Acid metabolism
p. 57 of 65
Courtesy
dnareplication.info
29 April 2008
How close to instability is it?



Pretty close.
Heating DNA makes it melt: fig. 19.17
The more GC pairs, the harder it is to
melt


Weaker stacking interactions in A-T
One more H-bond per GC than per AT
Amino Acid metabolism
p. 58 of 65
29 April 2008
iClicker quiz

3. What positions of a pair of aromatic
rings leads to stabilizing interactions?





(a) Parallel to one another
(b) Perpendicular to one another
(c) At a 45º angle to one another
(d) Both (a) and (b)
(e) All three: (a), (b), and ( c)
Amino Acid metabolism
p. 59 of 65
29 April 2008
Final iClicker question!

4. Which has the highest molecular mass
among the compounds listed?





(a) cytidylate
(b) thymidylate
(c) adenylate
(d) adenosine triphosphate
(e) they’re all the same MW
Amino Acid metabolism
p. 60 of 65
29 April 2008
Base composition for DNA


As noted, [A]=[T], [C]=[G] because of
base pairing
[A]/[C] etc. not governed by base pairing
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Can vary considerably (table 19.2)
E.coli : [A], [C] about equal
Mycobacterium tuberculosis: [C] > 2*[A]
Mammals: [C] < 0.74*[A]
Amino Acid metabolism
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Supercoiling
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Refers to levels of organization of DNA
beyond the immediate double-helix
We describe circular DNA as relaxed if
the closed double helix could lie flat
It’s underwound or overwound if the ends
are broken, twisted, and rejoined.
Supercoils restore 10.4 bp/turn relation
upon rejoining: see fig. 19.19.
Amino Acid metabolism
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Supercoiling
and flat DNA
Diagram courtesy SIU Carbondale
Amino Acid metabolism
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Ribonucleic acid
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We’re done with DNA for the moment.
Let’s discuss RNA.
RNA is generally, but not always, singlestranded
The regions where localized base-pairing
occurs (local double-stranded regions)
often are of functional significance
Amino Acid metabolism
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RNA physics & chemistry
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RNA molecules vary widely in size, from a few
bases in length up to 10000s of bases
There are several types of RNA found in cells
Type
%%turn- Size,
RNA over
by
mRNA 3
25 50-104
tRNA 15
21 55-90
rRNA 80
50 102-104
sRNA
2
4 30-103
Amino Acid metabolism
Partly
DS?
no
yes
no
?
Role
protein template
aa activation
transl. catalysis &
scaffolding
various
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