CHAPER 24

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CHAPER 24 AMINO ACIDS
AND PROTEINS
24.1 INTRODUCTION
Of the three groups of biopolymers, protein have
the most diverse function. Most of its molecular
weights are much larger. Their shaps cover a
range from the globular protein to the helical
coils of a akeratin. But all proteins have
common features.
Proteins are polyamides and their monomeric units
are about 20 different a-amino acids.
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Primary structure the exact sequence of the
different a-amino acids
along the protein chain.
Second and tertiary structure the folding of the
polyamide chain
which give rise to higher levels
of
complexity.
Although hydrolysis of natural occurring proteins
may yield as many as 22 different amino acids,
the amino acids have an important structural
feature in common.
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24.2 AMINO ACIDS
24.2A STRUCTURES AND NAMES
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The conversion of cysteine to cystine requires
addition comment. it can be reversed by mild
reducing agents.
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24.2B ESSENTIAL AMINO ACIDS
For adult humans there are eight essential amino
acids. These are designated with the superscript
e in above table.
24.2C AMINO ACIDS AS DIPOLAR IONS
Since amino acids contain both a basic group
(-NH2) and an acidic group (-COOH) , they are
amphoteric.
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In strongly basic solutions all amino present as
anions, in acidic solutions they are present as
cations. At some intermediate Ph, called
isoelectric point(pI) the concentration of the
dipolar ion is at its maximum and the
concentrations of the anions and cations are
equal.
As the acidity reaches pH 2.3, one half of the
cationic form will be converted to the dipolar
ion. As the pH increase to2.3- 9.7
the predominant form will be the dipolar ion.
When pH rise to 9.7,the dipolar ion will be
half-converted to the anionic form. As pH
approached to 14,the anionic form becomes
predominant form.
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If the side chain of an amino acid contains an
extra acidic or basic group, then the equilibria
are more complex.
The isoelectric point (pI) of an amino such as
the alanine is the average of pKa1 and pKa2.
24.3 LABORATORY SYNTHESIS OF a-AMINO ACIDS
A variety of methods have been developed for the
laboratory synthesis of a-amino acids. We shall
describe here three general methods.
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24.3A DIRECT AMMONOLYSIS OF AN a-HALO ACID
This method is probably used least often because
yields tend to be poor.
24.3B FROM POTASSIUM PHTHALIMIDE
This method is a modification of the Gabriel
synthesis of amines. the yields are usually high
and the products are easily purified.
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24.3C THE STRECKER SYNTHESIS
Treating an aldehyde with ammonia and hydrogen
cyanide produces an a-amino nitrile. Hydrolysis
of the nitrile group of the a-amino nitrile
converts the latter to an a-amino acid.
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Mechanism of the first step
24.3D RESOLUTION OF DL-AMINO ACIDS
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One interesting method for resolving amino acids
is based on the use of enzymes called deacylases.
24.3E STEREOSELECTIVE SYNTHESIS OF AMINO
ACIDS
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Producing only the naturally occurring L-amino
acid has been realized through the use of chiral
hydrogenation catalysts from transition metals.
One of which is called (R)-prophos.
Hydrolysis of N-acetyl group under this chiral
rhodium complex yields L-alanine. Because the
hydrogenation catalyst is chiral, it transfers
its hydrogen atoms in a stereoselective way. This
type of reaction is called asymmetric synthesis.
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24.4 ANALYSIS OF AMINO ACID MIXTURES
Enzymes can cause a-amino acids to polymerize
through the elimination of water
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The CO-NH- linkage between the amino acids is
called a peptide bond. Amino acid when joined in
this way, are called amino acid residues. The
polymers that contains 2,3, a few, or many amino
acid residues are called dipeptides, tripeptides,
oligopeptides, and polypeptides, respectively.
Polypeptides are linear polymers. The free
group and the free group are called
the N-terminal and the C-terminal Residues
respectively.
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The automatic amino acid analyzers are based on
the use of insoluble polymers containing
sulfonate groups, called cation-exchange resins.
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If the mixture of amino acids pass through a
column which is washed with a buffered solution
at a given pH, The individual amino acids will
move down the column at different rates and
ultimately separated.
24.5 AMINO ACID SEQUENCE OF POLYPEPTIDES AND
PROTEINS
The different amino acid sequences of a protein
which compose of 20 different amino acids in a
single chain of 100 residues are Amazing large.
They are
The methods of determining the amino acid
sequence include Terminal residue analysis,
partial hydrolysis and so on.
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24.5A TERMINAL RESIDUE ANALYSIS
One very useful method for determining the
N-terminal amino acid residue, called the Sanger
method, is based on the use of 2,4- dinitrofluorob
enzene (DNFB).
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A second method of N-terminal analysis is the
Edman degradation. This method offers an
advantage over the Sanger method in that it
Moves the N-terminal residue and leaves the
remainder of the peptide Chain intact.
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C-terminal residues can identified through the
use of digestive enzymes called
carboxypeptidases. These enzymes
specifically catalyze the hydrolysis of the amide
bond of the amino acid residue containing a free
COOH group, liberating it as a free amino acid.
24.5B PARTIAL HYDROLYSIS
Break the polypeptide chain into small fragments,
then examine the structure of these smaller
fragments to determine the original polypeptide.
For example
We are given a pentapeptide known to contain
valine(two residues), lucine, Histidine, and
phenylalanine. Then the molecular formular
Val2, Leu, His, Phe
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By using DNFB and carboxypeptidase we discover
that valine and leucine are the N-terminal and
C-terminal, respectively.
Val ( Val, His, Phe) Leu
We then subject the pentapeptide to partial acid
hydrolysis and obtain the following dipeptides.
ValHis HisVal ValPhe PheLeu
The points of overlap of the dipeptides tell us
that the original Pentapeptide must have been the
following
Val His Val Phe Leu
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24.6 PRIMARY ATRUCTURES OF POLYPEPTIDES
AND PROTEINS
The covalent structure of a protein or
polypeptide is called primary structure. Chemists
have had remarkable success in determining the
primary structure.
24.6A OXYTOCIN AND VASOPRESSIN
Oxytocin and vasopressin are two rather
polypeptides with strikingly similar structures.
But these two polypeptides have quite different
physiological effects.
Oxytocin occurs only in the female of a species
and stimulates uterine contraction during
childbirth. Vasopressin occurs in male and
female. Its major function is as an antidiuretic.
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24.6B INSULIN
Insulin, a hormone secreted by the pancreas,
regulates glucose metabolism.
Bocine insulin has a total of 51 amino acid
residues in two poly- peptide chains, called A
and B chains. These chains are joined by two
disulfide linkage.
Human insulin differs from bovine insulin at only
three amino acids residues. Insulin from most
mammals has a similar structure.
24.6C OTHER POLYPEPTIDES AND PROTEINS
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• Successful sequential analyses have now been
  achieved with
• hundreds of other polypeptides and proteins
  including the
• following
• Bovine ribonuclease.
• human hemoglobin.
• bovine trypsinogen and chymotrypsinogen.
• gamma globulin.

24.7 POLYPEPTIDE AND PROTEIN SYNTHESIS
We must first activate the carboxyl group of an
acid by converting it to an anhydride or acid
chloride and then allow it react with an amine.
But when both the acid group and the amino group
are present in the same molecular, the problem
becomes more complicate.
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24.7A PROTECTING GROUPS
We must protect the amino group by converting
it to some other group of low nucleophilicity-one
that will not react with a reactive acyl
derivative. Then remove the protecting group.
The reagents are benzyl chloroformate and
di-tert-butyl carbonate
Both reagents react with the following amino
group to form derivatives that are unreactive
toward further acylation.
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Remove of the benzyl group with hydrogen and a
catalyst depends on the fact that benzyl-oxygen
bonds are weaker and are subject to
hydrogenolysis at low temperatures.
24.7B ACTIVATION OF THE CARBOXYL GROUP
A much better method is to convert the carboxyl
group of the protected amino acid to a mixed
anhydride using ethyl chloro- Formate.
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The mixed anhydride can be used to acylate
another amino acid and form a peptide linkage.
Dicyclohexylcarbodiimide can also be used to
activate the carboxyl group of an amino acid.
24.7C PEPTIDE SYNTHESIS
The principle involve here can,of course, be
extended to the synthesis of much longer
polypeptide chains.
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24.7D AUTOMATED PEPTIDE SYNTHESIS
The Merrifield method for automated synthesis
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24.8 SECONDARY AND TERTIARY STUCTURES
OF PROTEINS
24.8A SECONDARY STRUCTURE
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The secondary structure of a protein is defined
by the local Confor- mation of its polypeptide
backbone. These local conformation have come to
be specified terms of regular folding patterns.
Polypeptide chain of a natural protein can
interact with itself two major ways through
formation of a ß-pleated sheet and an a helix.
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Fully extended polypeptide chains could
conceivably form a flat- sheet structure (above).
Slight rotation of bonds can transform a
flat-sheet structure into the ß-pleated sheet or
ß configuration.
The a helix structure is a right-handed helix
with 3.6amino acid residues per turn in
naturally occurring. It is the predominant
structure of the polypeptide.a
Helices and pleated sheets account for only about
one half of the average globular protein. The
remaining polypeptide segments have what is
called a coil or loop conformation.
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24.8B TERTIARY STRUCTURE
The tertiary structure of a protein is its
three-dimensional shape that arises from further
foldings of its polypeptide chains, foldings
superimposed on the coils of the a helixes.
24.9 INTRODUCTION TO ENZYMES
Enzymes have the ability to bring about vast
increases in the rates of reaction. Enzymes also
show remarkable specificity for their reactants
and for their products.
The enzyme and the substrate combine to form an
enzyme-substrate complex.
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Almost all enzymes are proteins. Reactions
catalyzed by enzymes are completely
stereospecific, and this specificity comes from
the way enzymes bind their substrates.
Some enzymes will accept only one compound as its
substrate, others will accept a range of
compounds with similar groups.
Inhibitor a compound that can alter the activity
of an enzyme. competitive inhibitor a compound
that competes directly with
the substrate for the active
site.
Some enzymes require the presence of a cofactor.
Others may require the presence of an organic
molecule called a coenzyme.
Many of the water-soluble vitamins are the
precursors of coenzymes.
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24.10 LYSOZYME MODE OF ACTION OF AN ENZYME
Lysozyme is made up of 129 amino acid residues.
Three short segments of the chain between
residues 5-15, 24-34, 88-96 have the structure of
an a helix the residues between 41-45, and 50-54
form pleated sheets and a hairpin turn occurs
at residues 46-49. The remaining polypeptide
segments of lysozyme have a coil or loop
conformation.
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Lysozymes substrate is a polysaccharide of amino
sugar that makes up part of the bacterial cell
wall.
24.11 SERINE PROTEASES
Serine proteases the digestive enzymes secreted
by the pancreas into
the small intestines to catalyze the hydrolysis
of peptide bonds.
The digestive enzymes includes chymotrypsin,
trypsin, and elastin.
The catalytic triad of chymotrypsin cause
cleavage of a peptide bond by acylation of the
serine residue 195 of chymotrysin. Near the
active site is a hydrophobic binding site that
accommodates nonpolar side chains of the protein.
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Regeneration of the active site of chymotrypsin.
Water causes hydrolysis of the acyl-serine bond.
Compounds such as diisopropylphosphofluoridate
(DIPF) that irreversibly inhibit serine
proteases. It has been shown that they do this
by reacting only with Ser 195.
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24.12 HEMOGLOBIN A CONJUGATED PROTEIN
Hemoglobin a protein can carry oxygen.
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The iron of the heme group is in the 2 oxidation
state and it forms a coordinate bond to a
nitrogen of the imidazole group of histidine of
the polypeptide chain. This leaves one valence of
the ferrous ion combine with oxygen as follows
When the heme combing with oxygen the ferrous ion
does not become readily oxidized to the ferric
state.