Amino Acid, Peptide,

Protein
Harliansyah, Ph.D
Head Dept of Biochemistry,
FKUY
2010

Email : ianshr2001@yahoo.com
A. Physical Properties

1. Solubility - soluble in water and insoluble in
organic solvents

2. Melting Points - melt at higher temperatures >
often 200C

3. Taste
sweet (Gly, Ala, Val)
tasteless (Leu)
bitter (Arg, Ile)
Sodium Glutamate
salt of Glutamic Acid flavoring agent
4. Optical Properties
- Assymetric a carbon atom is
attached to 4
different groups

exhibiting optical isomerism


All AA except Glycine possess
optical isomers due to
asymmetric -carbon atom

Some AA (Isoleucine, Threonine)
2
nd
asymmetric carbon

D- and L- forms of AA based on
the structure of glyceraldehyde


5. Amino acids as ampholytes

can donate a proton or accept a
proton

AA contain both acidic (-COOH)
and basic (-NH2) groups
Zwitterion or dipolar ion:
Zwitter
from German word means
hybrid

Zwitter ion (or dipolar ion)
a hybrid molecule containing (+) and (-) ionic
groups
AA rarely exist in a neutral form with free carboxylic (-COOH) and
free Amino (-NH2) groups
Strongly acidic pH (low pH) AA (+)
charged (cation)
Strongly alkaline pH (high pH) AA (-)
charged (anion)
Each AA has a characteristic pH (e.g. Leucine, pH 6.0), at which
it carries both (+) and (-) charges and exist as zwitterion
Chemical Properties

General Reactions mostly due to the 2
functional groups

Reactions due to - COOH group

1. AA from salts (-COONa) with bases
and esters (-COOR) with alcohols

2. Decarboxylation
- AA undergo decarboxylation to produce corresponding amines
3. Reaction with Ammonia

- the carboxyl group of dicarboxylic AA reacts with NH3 to form amide

Asparatic Acid + NH3 Asparagine
Glutamic Acid + NH3 Glutamine
Reactions due to -NH2 group
4. The Amino groups behave as bases and
combine with acids (e.g. HCl) to form
salts (-NH3 + Cl)

5. Reaction with NINHYDRIN (Ruhemanns blue purple)

6. Colour reactions of Amino Acids
- AA can be identified by specific colour reactions
Color Reactions of proteins / AA
Reaction Specific group or AA
1. Biuret Reaction Two peptide linkages
2. Ninhydrin Reaction -Amino acids
3. Hopkins Cole Reaction Indole ring of aromatic AA (Trp)
4. Millions reaction Phenolic Group (Tyr)
5. Xanthoprotein Reaction Aromatic ring (Phen, Tyr,Trp)
Millons Test
6. Sakaguchi Reaction Guanidino Group (Arg)
7. Nitroprusside Reaction Sulfhydryl groups (Cys)
8. Paulys test Imidazole ring (His)
9. Sulfur test Sulfhydryl groups (Cys)
10. Folin Coicalteaus Phenolic groups test (Tyr)

Fohls Reaction
7. Transamination
- important reaction in AA metabolism
- transfer of an amino group from an amino acid to a keto acid to form a new
AA
8. Oxidative deamination
- AA undergo oxidative deamination to liberate free ammonia
Characteristics of Peptide Bonds
1. Rigid
2. Planar
3. Partial double bond in character
4. Generally exists in transconfiguration
5. Both _C = O and _NH
2
groups of peptide
bonds are polar
6. Involved in hydrogen bond formation
Writing of Peptide Structures
The peptide chains are written with the free
Amino end (N terminal residue) at the left,
and the free carboxyl end (C terminal residue)
at the right.
The AA sequence is read from N terminal end
to C terminal end
Incidentally the protein biosynthesis also starts
from the N terminal Acid.
Naming of Peptides
For naming peptides, the AA suffixes ine
(glycine), - an (tryptophan) ate (glutamate) are
changed to yl with the exception of C
terminal AA.


A tripeptide composed of an N terminal
glutamate, a cysteine and a C terminal glycine
is called:
glutamyl cysteinyl - glycine
Polypeptide Chains
- The linking together of many AA by peptide
bonds produces polypeptide chains.
a.Residues - AA, when in polypeptide
chains, are customarily referred to as
residues.
b.Large peptide chains - protein
polypeptide chains are typically more
than 100 AA residues long. The
backbone of the chain is a recurring
sequence:
alpha - carbon carbonyl carbon -
alpha - amino N - alpha - carbon
Bovine Insulin: the first sequenced protein
Human: Thr-Ser-Ile
Cow: Ala-Ser-Val
Pig: Thr-Ser-Ile

Chiken: His-Asn-Thr
The role of side chain in the
shape of proteins
Where is water?
Hydrophobic
Hydrophilic
Amino acid substitution in proteins from different species
Conservative
Substitution of an amino acid by another
amino acid of similar polarity
(Val for Ile in position 10 of insulin)
Non conservative
Substitution involving replacement
of an amino acid by another of
different polarity
(sickle cell anemia, 6th position of hemoglobin
replace from a glutamic acid to a valine induce
precipitation of hemoglobin in red blood cells)
Invariant residues Amino acid found at the same position in
different species
(critical for for the sructure or function of the protein)
Biuret Reaction

Stabilized by hydrogen bonds
H- bonds are between CO and NH
groups of peptide backbone
H-bonds are either intra- or inter-
molecular
3 types : a-helix, b-sheet and triple-helix
Two type of b
Sheet structures
An anti paralellel
b sheet
A paralellel
b sheet
Triple helix of Collagen
Peptides of Physiologic Importance
1. Glutamine (Glutathione)
- a tripeptide composed
of 3 AA
- gamma glutamyl
cysteinyl glycine
- wildly distributed in
nature
- exists in reduced or
oxidized states

Functions:
a) As a coenzyme for certain enzymes as
prostaglandin PGE
2
synthase
glycoxylase
b) Prevents the oxidation of sulfhydryl groups of several proteins to
disulfide groups
c) In association with glutathione reductase participates in the
formation of correct disulfide bonds in several protiens
d) In erythrocytes
- maintains RBC membrane structure and
integrity
- protects hemoglobin from getting oxidized by agents such
as H
2
O
2
e) Involved in the transport of AA in the
intestine and kidney tubules via delta
glutamyl cycle or Meister cycle

f) Involved in the detoxification process

g) Toxic amounts of peroxidases and free
radicals produced in the cells are scavanged
by glutathione peroxidase ( a selenium
containing enzyme).
2. Thyrotropin Releasing Hormone (TRH)
- a tripeptide secreted by hypothalamus
Function:
Stimulate pituitary gland to release thyrotropic hormone

3. Oxytocin
- contains 9 AA (nonapeptide)
- hormone secreted by posterior
pituitary gland
Function:
Stimulate contraction of the uterus muscle during delivery

Stimulate contraction of muscle in breasts for milk ejection
Oxytocin and vasopressin are two peptide hormones with
very similar structure, but with very different biological
activities.

Interestingly, their structures only differ by one amino acid
residue (the hydrophobic LEU number 8 in oxytocin is replaced
by a hydrophilic ARG residue in vasopressin).

Oxytocin is a potent stimulator of uterine smooth muscle, and
also stimulates lactation.

Vasopressin, also know as antidiuretic hormone (ADH), has
no effect on uterine smooth muscle, but causes reabsorbtion
of water by the kidney, thus increasing blood pressure.
4. Vasopressin (ADH antidiuretic hormone)
- also a nonapeptide
- produced by posterior pituitary gland
Function:
Stimulates kidneys to retain water and thus
increases the blood pressure

5. Angiotensins
- Angiotensin 1 a decapeptide (10AA) which is
converted to angiotensin II (8AA)
Function:
For the release of aldosterone from adrenal gland
6. Methionine Enkephalin
- a pentapeptide found in the brain and has opiate
like function.
Function:
It inhibits the sense of a pain.

7. Bradykinin and Kallidin
- nona and decapeptides respectively
- produced from plasma proteins by
snake venom enzymes
Function:
Powerful vasodilators
8. Peptide Antibiotics
- Antibiotics such as Gramicidin, Bacitracin,
tyrocidin and Actinomysin
peptide in nature

9. Dipeptide aspartame
- Consists of aspartate and phenylalanine
- acts as Sweetener ~ used by diabetic patients

10. Gastrointestinal Hormones
- Gastrin, Secretin & etc.
- gastrointestinal peptides serving as
hormones

- they differ in their
physicochemical properties
which ultimately determine
the characteristics of proteins
The Proteins speak:
We are the basis of structure
and function of life;
Composed of twenty amino acids,
the building blocks;
Organized into primary, secondary, tertiary
and quaternary structure;
Classified as simple, conjugated
and derived proteins.
- 4 different forces stabilize the tertiary structure of globular protein
i. Hydrogen bonding between R groups of residues in
adjacent loops of the chain
ii. Ionic attraction between oppositely charged R groups
iii. Hydrophobic interactions
iv. Covalent cross-linkages (via intrachain cystein residues)
Function of proteins
Proteins
are the most
important buffers in
the body.
Enzymatic catalysis
Transport and storage (the protein hemoglobin,
albumins)
Coordinated motion (actin and myosin).

Mechanical support (collagen).

Immune protection (antibodies)

Generation and transmission of nerve impulses
- some amino acids act as neurotransmitters,
receptors for neurotransmitters, drugs, etc. are
protein in nature. (the acetylcholine receptor),

Control of growth and differentiation -
transcription factors
Hormones
growth factors ( insulin or thyroid stimulating
hormone)
Why?
(a) Protein molecules
possess basic and
acidic groups which act
as H+ acceptors or
donors respectively if
H+ is added or removed.

(a) Proteins are the most important buffers in the body. They are mainly
intracellular and include haemoglobin.

(b) The plasma proteins are buffers but the absolute amount is small compared
to intracellular protein.

(c) Protein molecules possess basic and acidic groups which act as H+
acceptors or donors respectively if H+ is added or removed.
Many proteins (thousands!) present in blood plasma
Proteins contain weakly acidic (glutamate, aspartate) and basic
(lysine, arginine, histidine) side chains (or R groups)
At neutral pH, only histidine residues (containing imidazole R
group with pK
a
~ 6.0) in proteins can act as a buffer component
Haemoglobin with 38 histidine/tetramer is a good buffer
N-terminal groups of proteins (pK
a
~ 8.0) can also act as a buffer
component
Classes of protein
1 - Structural protein : Keratin, Collagen (give support to cells)
2 - Dynamic protein : Hormone, enzyme (for catalytic purpose)

- Based on the structure, protein can be divided to :
* Fibrin : Blood clotting
* Fibrous : Myosin (from muscle)
* Globular : Half sphere form/structure eg. Enzyme
- Size : Varied - depending on functions
- 1 amino acid = 110 Daltons
- Most protein are highly folded
Fibrous and Globular Proteins
- There are also protein which its polypeptide chains are tightly
folded into a spherical or globular shape
Example of globular protein :
Lysozyme molecule with its tightly
bound polysaccharide substrate
(color)
Enzyme Catalysis
Transport and storage - small molecules are often carried by proteins in the physiological setting
(for example, the protein hemoglobin is responsible for the transport of oxygen to tissues). Many drug
molecules are partially bound to serum albumins in the plasma.
3-dimensional structure of hemoglobin.
The four subunits are shown in red and
yellow, and the heme groups in green.
The binding of oxygen is affected by molecules such as carbon
monoxide (CO) (for example from tobacco smoking, cars and
furnaces).

CO competes with oxygen at the heme binding site. Hemoglobin
binding affinity for CO is 200 times greater than its affinity for
oxygen, meaning that small amounts of CO dramatically reduces
hemoglobin's ability to transport oxygen. When hemoglobin
combines with CO, it forms a very bright red compound called
carboxyhemoglobin.

When inspired air contains CO levels as low as 0.02%, headache
and nausea occur; if the CO concentration is increased to 0.1%,
unconsciousness will follow. In heavy smokers, up to 20% of the
oxygen-active sites can be blocked by CO.
Coordinated motion - muscle is mostly protein, and muscle contraction is mediated by the sliding
motion of two protein filaments, actin and myosin.
Platelet before activation Activated platelet
Activated platelet
at a later stage than C)
Platelet activation is a controlled
sequence of actin filament:

Severing
Uncapping
Elongating
Cross linking

That creates a dramatic shape change
in the platelet
Mechanical support - skin and bone are strengthened by the protein collagen.
Abnormal collagen synthesis or
structure causes dysfunction of

cardiovascular organs,
bone,
skin,
joints
eyes

Refer to Devlin
Clinical correlation 3.4 p121
Immune protection - antibodies are protein structures that are responsible for reacting with specific
foreign substances in the body.
Generation and transmission of nerve impulses -

Some amino acids act as neurotransmitters, which transmit electrical signals from one nerve cell to another. In
addition, receptors for neurotransmitters, drugs, etc. are protein in nature.

An example of this is the acetylcholine receptor, which is a protein structure that is embedded in postsynaptic
neurons.

GABA:
gamma Amino butyric acid
Synthesised from glutamate

GABA acts at inhibitory synapses in the
brain. GABA acts by binding to specific
receptors in the plasma membrane of both
pre- and postsynaptic neurons.
Neurotransmetter
Membrane transport proteins
Protein degradation:
Disease
Example:
Neurodegenerative
diseases
Disease and protein folding:
Digestion of Proteins
1. Mouth - food is chewed
2. Stomach:
Gastrin - triggers chief cells to release HCl and
pepsinogen
3. Small intestines:
Secretin-stim. release of pancreatic juices
Activation of proteolytic enzymes:
Trypsinogen trypsin
Chymotrypsinogen chymotrypsin
Proelastase elastase
Endo & exopeptidases activated
Products of enzyme action = amino acids
Dipeptides & small peptides AA
Greater amount of AA in portal blood is in the form of
alanine
STOMACH
Protein digestion starts in the stomach
dietary proteins become denatured by gastric acid
important for protein digestion because proteins are
poor substrates for proteases

ACID ENVIRONMENT is required for action of pepsin
PEPSIN
A protease that works optimally at pH 2
a carboxyl protease
acts mostly as endopeptidase
does not cleave at random
prefers peptide bonds formed by amino group of
aromatic AA
major breakdown products are not free AA but a
mixture of oligopeptides known as peptones
INTESTINE
As acidic stomach contents reach duodenum
rapidly neutralized by HCO
3
in pancreatic secretions.

Proteolytic enzymes from pancreas include:
1. Trypsin
a serine protease
endopeptidase
specific for CO side of basic AA
2. Chymotrypsin
a serine protease
endopeptidase
specific for CO side of hydrophobic AA
1 & 2 = degrade peptones to smaller peptides

3. Carboxypeptidase A:
hydrophobic AA at C terminal

4. Carboxypeptidase B
basic AA at C terminus