Chapter 3

Amino Acids
and
Peptides

1
 Amino Acids
• Amino acid: a compound that contains both an
amino group and a carboxyl group
• -Amino acid: an amino acid in which the
amino group is on the carbon adjacent to the
carboxyl group
• although -amino acids are commonly written
in the unionized form, they are more properly
written in the zwitterion (internal salt) form
O O
R- CH-COH R- CH-CO -
N H2 N H3 +
unionized zwitterion
form

2
 Chirality of Amino Acids
• With the exception of glycine, all protein-derived amino
acids have at least one stereocenter (the -carbon) and
are chiral
• the vast majority of -amino acids have the L-
configuration at the -carbon

COO- COO-
H N H3 + +
H3 N H
CH3 CH3
D-Alanine L-Alanine
(Fischer projections)

3
 Chirality of Amino Acids
• Comparison of the stereochemistry of alanine and
glyceraldehyde (Fischer projection formulas)
the naturally
occurring form

COO- COO-
H N H3 + +
H3 N H
CH3 CH3
D-Alanine L-Alanine

CHO CHO
the naturally
H OH HO H
occurring form
CH2 OH CH2 OH
D-Glyceraldehyde L-Glyceraldehyde

4
 20 Protein-Derived AA
Nonpolar side chains (predominant form at pH 7.0)
glycine (gly, G) phenylalanine (phe, F)
H-
alanine (ala, A)
CH3 -
tryptophan (trp, W)
valine (val, V)
( CH3 ) 2 CH
N
leucine (leu, L)
H
( CH3 ) 2 CHCH 2 -
proline (Pro, P)
isoleucine (ile, I)
CH3 CH2 CH( CH3 ) - +
N
methionine (met, M) H H
CH3 S CH2 CH2 -

5
 20 Protein-Derived AA
Polar side chains (predominant form at pH 7.0)
asparagine (asn, N) serine (ser, S)
O
H2 N CCH 2 - HOCH2 -

glutamine (glu, G) threonine (thr, T)

O OH
H2 N CCH 2 CH2 - CH3 CH-

6
 20 Protein-Derived AA
Acidic side chains (predominant form at pH 7.0)

aspartic acid (asp, D) glutamic acid (glu, E)

O O
- -
OCCH2 - OCCH2 CH2 -

cysteine (cys, C) tyrosine (tyr, Y)

HS CH 2 - HO CH2 -

7
 20 Protein-Derived AA
Basic side chains (predominant form at pH 7.0)

arginine (arg, R) histidine (his, H)

N H2 + N
H2 N CN HCH2 CH2 CH2 - CH2 -
N
lysine (lys, K) H
+
H3 N CH 2 CH 2 CH 2 CH 2 -

8
 20 Protein-Derived AA
• Note these structural features
1. All 20 are -amino acids
2. For 19 of the 20, the -amino group is primary; for
proline, it is secondary
3. With the exception of glycine, the -carbon of each is a
stereocenter
4. Isoleucine and threonine contain a second
stereocenter
5. The sulfhydryl group (pKa 8.3) of cysteine, the
imidazole group (pKa 6.0) of histidine, and the phenolic
hydroxyl (pKa 10.1) of phenylalanine are partially
ionized at pH 7.0, but the ionic form is not the major
form at this pH

9
 Uncommon Amino Acids
• Each example is derived from a common amino acid by
the modification shown in color
HO I I
OH
+
H3 N COO- HO O COO-
+
+ N COO-
N H3 N H3 +
H H I I
Hydroxylysine Hydroxyproline Thyroxine

• hydroxylysine and hydroxyproline are found only in a

few connective tissues such as collagen
• thyroxine is found only in the thyroid gland

10
 Ionization of Amino Acids
+1 charge 0 charge -1 charge
+ +
H3 N COOH H3 N COO- H2 N COO-
pK a = 2.34 pK a = 9.69

Isoelectric zwitterion

+2 charge +1 charge
+
H3 N COOH +
H3 N COO-
pK a = 1.82 pK a = 6.04
NH NH
+N +N
H H
0 charge -1 charge
+
H3 N COO- H2 N COO-
pK a = 9.17
NH NH
N N
Isoelectric zwitterion
11
 Titration of Amino Acids
Figure (a) Titration of alanine with NaOH

12
 Titration of Amino Acids
Figure (b) Titration of histidine with NaOH

13
 Acidity: -COOH Groups
• The average pKa of an -carboxyl group is 2.19, which
makes them considerably stronger acids than acetic acid
(pKa 4.76)
• the greater acidity of the amino acid carboxyl group is
due to the electron-withdrawing inductive effect of the
-NH3+ group
The ammonium ion has an
electron-withdrawing
inductive effect
pK a = 2.19 - +
RCHCOOH + H2 O RCHCOO + H3 O
+ +
N H3 N H3

14
 Acidity: -NH3+ groups

• The average value of pKa for an -NH3+ group is 9.47,

compared with a value of 10.76 for a 2° alkylammonium
ion
pK a = 9.47
- - +
RCHCOO + H2 O RCHCOO + H3 O
+
N H3 N H2

pK a = 10.76
+
CH3 CHCH 3 + H2 O CH3 CHCH 3 + H3 O
+
N H3 N H2

15
 Basicity: Guanidine Group
• The side chain of arginine is a considerably stronger base
than an aliphatic amine
• basicity of the guanido group is attributed to the large
resonance stabilization of the protonated form relative
to the neutral form +
:

N H2 N H2 N H2
+
:

:
RN H C RN H C RN H C
+ : N H2 : N H2
N H2 :

H2 O N H2
+
:

RN C + H3 O pK a = 12.48
: N H2

16
 Basicity: Imidazole Group
• The imidazole group on the side chain of histidine is a
heterocyclic
H + aromatic amine
H
N

:
N
- - H2 O
CH2 CHCOO
N + CH2 CHCOO
:

N +
N H3 +
H : H N H3

this lone pair is N

not a part of the - +
CH2 CHCOO + H3 O pK a 6.04
:

aromatic sextet; it is N
+
the proton acceptor H N H3

17
 Ionization vs pH
• Given the value of pKa of each functional group, we can
calculate the ratio of each acid to its conjugate base as a
function of pH
• Consider the ionization of an -COOH
pK a = 2.00
- +
COOH + H2 O COO + H3 O

• writing the acid ionization constant and rearranging

terms gives
[ H 3 O + ] [ -COO - ] [ -COO - ] Ka
Ka = or =
[ -COO H] [ -COO H] [ H3 O+ ]

18
 Ionization vs pH
• substituting the value of Ka (1 x 10-2) for the hydrogen
ion concentration
- at pH 7.0 (1.0 x 10-2-7) gives
[ -COO ] Ka 1.00 x 10
= = = 1.00 x 10 5
[ -COO H] [ H3 O+ ] 1.00 x 10-7

• at pH 7.0, the -carboxyl group is virtually 100% in the

ionized or conjugate base form, and has a net charge
of -1
• we can repeat this calculation at any pH and determine
the ratio of [-COO-] to [-COOH] and the net charge on
the -carboxyl at that pH

19
 Ionization vs pH
• We can also calculate the ratio of acid to conjugate base
for an -NH3+ group; for this calculation, assume a value
10.0 for pKa
+ pK a = 10.00
N H3 + H2 O N H2 + H3 O +

• writing the acid ionization constant and rearranging

gives [ -NH 2 ] Ka
=
+
[ -NH 3 ] [H 3 O+ ]

20
 Ionization vs pH
• substituting values for Ka of an -NH3+ group and the
hydrogen ion concentration at pH 7.0 gives
[ -NH 2 ] Ka 1.00 x 10-10
+
= + = = 1.00 x 10 -3
[ -NH 3 ] [H 3 O ] 1.00 x 10-7

• at pH 7.0, the ratio of -NH2 to -NH3 + is approximately

1 to 1000
• at this pH, an -amino group is 99.9% in the acid or
protonated form and has a charge of +1

21
 Henderson-Hasselbalch
• We have calculated the ratio of acid to conjugate base for
an -carboxyl group and an -amino group at pH 7.0
• We can do this for any weak acid and its conjugate base
at any pH using the Henderson-Hasselbalch equation

[conjugate base]
pH = pK a + log
[weak acid]

22
 Henderson-Hasselbalch
• using the Henderson-Hasselbalch equation, we can
calculate the percent of charged or uncharged form
present and the net charge on serine at pH 3.0, 7.0, and
10.0
100% 86% 99% 100% 88% 100%
O O O
+ + - -
H3 N- CH-C- OH H3 N- CH-C- O H2 N- CH-C- O
CH2 OH CH2 OH CH2 OH
pH 3.0 pH 7.0 pH 10.0
Net charge +1 Net charge 0 Net charge -1

23
 Isoelectric pH
• Isoelectric pH, pI: the pH at which the majority of
molecules of a compound in solution have no net charge
• the pI for glycine, for example, falls midway between
the pKa values for the carboxyl and amino groups

pI = 1 ( p Ka COOH + p Ka N H3 + )
2

= 1 (2.35 + 9.78) = 6.06

2

• given in the following tables are isoelectric pH values

for the 20 protein-derived amino acids

24
 Isoelectric pH (pI)
Table 3.2 pK a and pI of -amino acids
Nonpolar & pK a of pK a of pK a of
polar side chains COOH N H3 + Side Chain pI
alanine 2.34 9.69 ---- 6.02
asparagine 2.02 8.80 ---- 5.41
glutamine 2.17 9.13 ---- 5.65
glycine 2.34 9.60 ---- 5.97
isoleucine 2.36 9.68 ---- 6.02
leucine 2.36 9.68 ---- 6.02
methionine 2.28 9.21 ---- 5.74
phenylalanine 1.83 9.13 ---- 5.48
proline 1.99 10.60 ---- 6.30
serine 2.21 9.15 ---- 5.68
threonine 2.63 10.43 ---- 6.53
tryptophan 2.38 9.39 ---- 5.89
valine 2.32 9.62 ---- 5.97
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 Isoelectric pH (pI)
Table 3.2 (cont'd)

Acidic pK a of pK a of pK a of
Side Chains COOH N H3 + Side Chain pI
aspartic acid 2.10 9.82 3.86 2.98
glutamic acid 2.10 9.47 4.07 3.08
cysteine 2.05 10.25 8.00 5.02
tyrosine 2.20 9.11 10.07 5.63

Basic pK a of pK a of pK a of
Side Chains COOH N H3 + Side Chain pI

arginine 2.01 9.04 12.48 10.76

histidine 1.77 9.18 6.10 7.64
lysine 2.18 8.95 10.53 9.74

26
 Electrophoresis
• Electrophoresis: the process of separating compounds
on the basis of their electric charge
• electrophoresis of amino acids can be carried out
using paper, starch, agar, certain plastics, and
cellulose acetate as solid supports
• in paper electrophoresis, a paper strip saturated with
an aqueous buffer of predetermined pH serves as a
bridge between two electrode vessels

27
 Electrophoresis （ Process)
• a sample of amino acids is applied as a spot (the
origin) on the solid support strip
• an electric potential is applied to the electrode vessels
and amino acids migrate toward the electrode with
charge opposite their own
• molecules with a high charge density move faster than
those with a low charge density
• molecules at their isoelectric point remain at the origin
• after separation is complete, the strip is dried and
developed to make the separated amino acids visible

28
 Polypeptides
• In 1902, Emil Fischer proposed that proteins are long
chains of amino acids joined by amide bonds to which he
gave the name peptide bonds
• Peptide bond: the special name given to the amide bond
between the -carboxyl group of one amino acid and the
-amino group of another

29
 Serylalanine (Ser-Ala)
peptide
bond
O HOH2 C H H
HOH2 C H + O
H3 N + N
+ O- + O- H3 N O-
H3 N
O H CH3 O H CH
3
Serine (Ser) Alanine (Ala) Serylalanine (Ser-Ala)

30
 Peptides
• peptide:
peptide the name given to a short polymer of amino
acids joined by peptide bonds; they are classified by
the number of amino acids in the chain
• dipeptide:
dipeptide a molecule containing two amino acids
joined by a peptide bond
• tripeptide:
tripeptide a molecule containing three amino acids
joined by peptide bonds
• polypeptide:
polypeptide a macromolecule containing many amino
acids joined by peptide bonds
• protein:
protein a biological macromolecule of molecular
weight 5000 g/mol or greater, consisting of one or
more polypeptide chains

31
 Geometry of Peptide Bond
• the four atoms of a peptide bond and the two alpha
carbons joined to it lie in a plane with bond angles of
120° about C and N
• to account for this geometry, Linus Pauling proposed
that a peptide bond is most accurately represented as
a hybrid of two contributing structures
• the hybrid has considerable C-N double bond
character and rotation about the peptide bond is
-
restricted :O

:
: :
:

C O C

+
C N
:

C N
C H C H

(1) (2)
32
 Writing Peptides
• By convention, peptides are written from the left,
beginning with the free -NH3+ group and ending with the
free -COO- group
• the repeat pattern, starting from the N-terminal amino
acid, is N ---> -carbon ---> carbonyl carbon etc.
peptide
bonds
S

O O C-terminal
+ H amino acid
H3 N N
N O-
H
N-terminal O N H2
amino acid OH
Ser-Met-Asn O

33
 Some Small Peptides
O O O
+ +
H3 N -CH- C-N H-CH-C-OCH3 H3 N -CH2 - CH 2 -C-N H- CH-COO -
CH2 CH2 CH2
COO- C6 H5
NH
L-Aspartyl-L-phenylalanine -Alanyl-L-histidine
methyl ester N
(Carnosine)
(Aspartame)

34
 Glutathione
N H3 + H O
- -
O N O - 2e oxidation
N
O O H O 2e- reduction
SH
Glutathione, GSH
(reduced form) N H3 + H O
-
O N O-
N
O O H O
S
A disulfide
S bond
O O H O
- N
O N O-
N H3 + H O
Glutathione, GS-SG
(oxidized form)

35
 Enkephalins
Leucine enkephalin
Tyr-Gly-Gly-Phe-Leu = Y-G-G-F-L

Methionine enkephalin
Tyr-Gly-Gly-Phe-Met = Y-G-G-F-M

36
 Oxytocin & Vasopressin
+ +
H3 N -Cy s - Ty r-Ile H3 N -Cy s - Ty r-Ph e
S Gln S Gln
S S
Cy s -A s n Cy s -A s n
O O
Pro -Le u - Gly -C- NH2 Pro -A rg - Gly- C-N H2
Oxytocin Vasopressin

37
End

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