Ionization of Amino Acids

• In amino acids, carboxyl group (-) and amino group (+) are
charged at neutral pH.

Dr. M. khalifeh
Ionic forms of amino acids

H R

C
COO-
H +
+
H3N H+
Zwitterion
H R pH 7 Net charge 0
H R

C C
+ COOH COO -
H3N H2N
pH 1 Net charge +1 pH 13 Net charge -1

Dr. M. khalifeh
Ionization of Amino Acids
• Remember, amino acids without charged groups on side chain
exist in neutral solution as isoelectric Zwitterion with no net
charge

Dr. M. khalifeh
Titration of Amino Acids (alanine)
• When an amino acid is titrated, the titration curve
represents the reaction of each functional group with
the hydroxide ion
PI= Isoelectric point
Is the point where the
molecule has no net charge

Dr. M. khalifeh
Titration of histidine with NaOH
Histidine has -1 charge
PI= PK2+PKa3 / 2

Histidine has +1 charge

Histidine has no charge

Histidine has +2 charge

Dr. M. khalifeh
Electrophoresis
• Alanine PI is 6 while at the same pH histidine has a
net charge of +1 (imidazole group is still protonated)
• So we can separate componds containing such a.a.
in electric field

Dr. M. khalifeh
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 to the electron-withdrawing inductive effect
of the -NH3+ group

Dr. M. khalifeh
Basicity:
• -NH3+ groups: The average value of pKa for an -
NH3+ group is 9.47
• 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
Dr. M. khalifeh
Basicity: Imidazole Group
• The imidazole group on the side chain of histidine is
a heterocyclic aromatic amine
H + 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

Dr. M. khalifeh
Polypeptides

• Peptide bond: the special name given to the amide

bond between the -carboxyl group of one amino
acid and the -amino group of another
• covalent linkage
• Also called amide
• Formation of this bond results in the release of 1 mol
of water per mol of peptide bond formed.
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)
Dr. M. khalifeh
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
• oligopeptides 20 or so amino acids joined by peptide
bonds
• polypeptide:
polypeptide a macromolecule containing many amino
acids joined by peptide bonds (more than hundred)
• protein:
protein a biological macromolecule of molecular
weight 5000 g/mol or greater, consisting of one or
more polypeptide chains
Dr. M. khalifeh
Geometry of Peptide Bond
• The resonance structure written with double bondin between C and N in the
peptide bond
• Which results in negative charge for the O and positive charge for the N
• The resonance structure prevents rotation arround the bond
• It is the results of the ability of the amide nitrogen to delocalize its lone pair
of electrons onto the carbonyl oxygen.
• The total charge must remain the same
• Not all bonds have resonance structure at all time thus not all peptide bonds
has double rigid bonds (approximately 40%)

: -
:O :
:

:O C C
+
C N
:

C N
C H C H

(1) (2)
Dr. M. khalifeh
Geometry of Peptide Bond
• the peptide bond is rigid, and
planar.
• Because of this resonance
• the C=O bond is actually
longer than normal carbonyl
bonds
• the N–C bond of the peptide
bond is shorter than the N–C
bond.
• Notice that the carbonyl oxygen
and amide hydrogen are in a
trans configuration, as opposed
to a cis configuration.
• This configuration is
energetically more favorable
because of possible steric
interactions in the other.Dr. M. khalifeh
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

Dr. M. khalifeh
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)

• artificial, non-saccharide • highly concentrated in

sweetener used as a sugar muscle and brain tissues
substitute • carnosine has a number of
antioxidant properties

Dr. M. khalifeh
Glutathione (tripeptide).
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
• It contains an unusual S
A disulfide
peptide linkage between the bond
S
amine group of cysteine and O O H O
the carboxyl group of the -
O N
N
O-
glutamate side chain. N H3 + H O
• an antioxidant, helps protect Glutathione, GS-SG
cells from reactive oxygen (oxidized form)
species such as free
radicals and peroxides
Dr. M. khalifeh
Glutathione (tripeptide).
• Glutathione is a scavenger for oxidizing agents
• Oxidation loss of e-
• oxidizing agents causes another agent to lose e-
• Reduced form  oxidized form
• Glutathione (2)  GSSG (Oxidized form) two GSH
(disulfide bond)

Dr. M. khalifeh
Enkephalins (penta p.p.)
Leucine enkephalin
Tyr-Gly-Gly-Phe-Leu = Y-G-G-F-L

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

• Differ only in C terminal a.a.

• Naturally occurring analgesics
• Aromatic sides of Y and F play functional role
• Morphine has similar 3D structure thus compete for
same receptor in brain and give same effect

Dr. M. khalifeh
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
• Peptides with cyclic structure (due to S-S)
• C terminal has amide group instead of carboxyl
group
• Act as hormones

Dr. M. khalifeh
Other cyclic peptides
• Gramicidin S and Tyrocidine A
• antibiotic (contain L and D a.a. forms)
• Contain the a.a. # 21 (intermidate form in metabolic
pathways)---- Ornithine (Orn)

Dr. M. khalifeh