AMINO ACIDS
-the identity/distinctive
PROTEINS
Most abundant macromolecules living
in cells
Great variety
Great diversity in biological function
Molecular instruments by which
genetic information is expressed
Gk protos : first or foremost
All are constructed from same
ubiquitous 20 AA
The 20 AA are joined in many
different combinations and sequences
Producing thousands of different
structures, different properties and
activities
Different products and enzymes,
hormones, antibodies, lens, feathers,
horns
AMINO ACIDS
Primary amines
Contain an amino group (NH3) group
connected to an alpha carbon and to
which other substituents are attached
carboxyl group (COOH)
Variable side chain R
H AA are L or D depending on the
A-amino acids because they have a
primary amino group (-NH2) and a
carboxylic acid group (-COOH)
attached to the alpha carbon
Amino acid common structure
consists of a central C atom which is
bonded to a:
-amino (-NH2)
- carboxyl group (-COOH)
-Hydrogen atom (-H)
-Side chain group (-R)
Variable side group (-R)
determines:
-its special properties
-size, shape, reactivity
>electric charge
>solubility in water
the optical activity of the isomer of
>size, structure
helps to dictate the folding of the
protein
Alpha Carbon atom
A hydrogen atom
Amino group (hence amino acid)
-the amino group (-NH2) accepts a
proton and becomes (-NH3+)
Carboxyl group (-COOH)
-this gives up a proton and is thus an
acid (hence amino acid)
-becomes dissociated (-COO-)
R group
No asymmetric carbon: GLYCINE is
not asymmetric
Alpha carbon is asymmetric (except
GLYCINE) chiral center
-tetrahedral bonding around the
alpha C
The 4 groups can occupy 2 different
arrangements in space, enantiomers
or stereoisomers (non-
superimposable mirror images)
Optically active
2 different arrangements of the
groups around the alpha carbon:
1. Enantiomers-mirror image
2. Stereoisomers
Non-superimposable
position of the amino group
Regular tetrahedron. It is
symmetrical and a mirror image is
no different from the original
The mirror images is not identical
The mirror image cannot be
superimposed on the original
handedness is called chirality and
the mirror images are called
enantiomers
If this compound is chiral and is
present as just one of the
enantiomers (mirror images), the
plane of the polarized light will be
rotated
The L and D convention for AA
configuration refers not to the optical
activity of the AA itself but rather to
involved in two carbon-nitrogen
glyceraldehyde from which that AA
can theoretically be synthesized (D-
glyceraldehyde is dextrorotary, L-
glyceraldehyde is levorotary)
Threonine and Isoleucine have 2
chiral carbons each, thus producing 4
possible stereoisomers each
L forms: -humans and most foods
are made up almost exclusively of L
form aminos
D form proteins are produced by
exotic sea-dwelling organisms such as
cone snails
-abundant components of the
peptidoglycan cell walls of bacteria
-D-serine may act as a
neurotransmitter in the brain
D-AA that occur naturally
-Free: D serine, D asp (brain tissue)
-D ala, D glu (cell wall of g+ bacteria)
-In peptides and antibiotics produced
by bacteria, fungi, reptiles
imidazole ring of histidine is aromatic
at all pH values
unprotonated imidazole is nucleophilic
(imidazole) and can serve as a
general base, while the protonated
form (imidazolium) can serve as a
general acid
acid form, the imidazolium ion (B), is
a resonance hybrid
imidazole Conjugate base (A or C)
Either of the two ring nitrogens can
release a proton (H+) to produce the
conjugate base form
Glycine (Gly, G) is the simplest and
smallest of all AA
-the only one which is not optically
active since it has a single H atom as
its side chain
Proline in which the R group makes
up part of a ring which also includes
the amino group and the alpha
carbon atom.
Since the amino group in proline is
bonds, it is a secondary amino group
Properties
Solubility:
-generally soluble in water and
insoluble in non-polar organic
solvents such as hydrocarbons
> Charged functional group: readily
soluble in polar solvents (H2O,
ethanol); insoluble in non-polar
solvents ( benzene, ether)
Colorless
-Aromatic AA absorb UV
MP
-amino acids are crystalline solids
with high melting points
Optical activity
-With chiral center: optically active
Ultraviolet absorption
-Tyr, Phe, Trp : absorb high wave-
length UV
> Try absorbs light at 280 nm
> Absorption bands arise from
interaction of radiation with electrons
on the aromatic rings
Acid base
-Amino and COOH groups
-pKa
pKa
- COOH : ~2.2
- Amino: ~9.4
At pH 7 the amino is protonated
NH3+ and carboxyl is in COO-
(conjugate base or carboxylate
form)
AA with single amino and single
carboxyl group will have net charge
of zero : Zwitterion; which are
electrically neutral
In an electric field : stationary
Zwitterions
- comes from the German word
zwitter meaning hybrid
- protonated ammonium group with a
(+) charge
- deprotonated carboxylate groups with
a (-) charge
- Net charge: NEUTRAL
- At certain compound specific pH:
Isoelectric point (pI)
1
 Isoelectric point (pI)
- pH at which AA has no net charge
(does not move in electric field)
-the ph at midway between the pK
values on either side of the Zwitterion
specie
pI of neutral AA ~6
-Acidic AA (very much<6)
-Basic AA (very much >6)
Physical properties are influenced by
ionic state of COOH and NH2 and
other ionizable group in the R
Each AA has either 2 or 3 pKa values
and these differ among different AA
At different pH AA have different net
charges
AA have both an amine and
carbocylic acid functional groups are
therefore both acid and base at the
same time
Zwitterions have minimal solubility at
their isoelectric point and AA are
often isolated by precipitation from
water after adjusting the pH to their
isoelectric point
-no mobility in electrophoresis
Amino acid in solution at isoelectric
pH are mainly dipolar ions
This is generally how AA exist at
cellular pH
Because of their amino and carboxyl
groups, proteins in solution resist
changes in acidity and alkalinity and
so are important biological buffers
Charged R (basic and acidic) stabilize
protein conformations thru ionic
interactions or salt bridges
Dissociation constant
amino group and the carboxyl group
of the amino acid have PKs of about
2.3 and 9.6 respectively
can exist in three general forms
PKs of AA
- ~9.6: amino group (mean for all
AA)
At pH < than this: NH3+
- ~2.3 carboxyl group (mean for the
20 AA: ~ 2.2)
At pH > 2.2: COO- a functional group that contains a C-N -basic polar: side chain contains
predominates double bond with the N atom an amino group
At pH between 2.2-9.4 the connected to an aryl or alkyl group
predominant state of the molecule: but not H Serine
COO- and NH3+ A mechanism used by enzymes to Threonine
-zwitterions catalyze rxn occurs through the Asparagine
Can be in 3 general forms: formation of imines (Schiff bases) Glutamine
1. Protonated form (dipolar with Cysteine
charge): Reversible oxidation of 2 Tyrosine
-they can dissociate cysteine side chain thiols to form
-NH3+ and COO- cysteine a. Non-polar
2. Unprotonated/ Neutral form: -R group is made up of only
-they cannot dissociate GRP 1 (nonpolar side chains) carbon and hydrogen
-NH2 and COOH Aliphatic HC side -non polar since there is very
3. Amino group: ~9.6 pH>NH3+ Alanine little polarity associated with
Carboxyl group: __________ Valine carbon-carbon and carbon-
Leucine hydrogen bonds (therefore
At low pH the amino and carboxyl Isoleucine hydrophobic)
groups will be protonated and the Proline aliphatic cyclic structure, N -Phe, Met, Ile, Leu, Val are
molecules will be in the acid form bonded to 2 C (secondary amine) very hydrophobic
COOH: ~2.2 Tryptophan : indole ring in R
As the pH is increased (beyond 2.2, Pheneylalanine (aromatic HC With Isoleucine
COOH dissociates) benzene ring) Phenylalanine
towards neutrality, the AA become Methionine R contains S (aliphatic ) Methionine
zwitterions having both negative and Glycine (no polar side chain) Leucine
positive charges Valine
AS the pH increase further (beyond GRP 2 (Polar R are neutral) Glycine
pK of aa) NH3 dissociates Serine (OH) Alanine
Threonine (OH) Proline
20 proteinogenic AA Asparagine (amide) Tryptophan
21st amino acid Selenocysteine Glutamine (amide)
-Found in some proteins Cysteine (SH) Heteroatoms
> Peroxidase reductase (ET rxns) Tyrosine (OH is phenol) -Sulfur
> Inserted into polypeptide during -Nitrogen
translation but not specified by codon GRP 3 (acidic Carboxyl in R) >Contribute very little polarity to the
22nd AA: L-Pyrollysine Glutamate side chain
- genetic code for pyrrolysine: in aspartate Heteroatoms in Methionine (sulfur)
Archae microbes named and tryptophan (Nitrogen)
Methanosarcina barkeri which GRP 4 (Basic side chains) -overall behavior of these AA: non-
produce methane or natural gas Histidine (imidazole) polar
Lysine Side chains which contain more polar
Reactions: Arginine (guanidino) functional groups such as acids,
Between AA and carboxyl groups amine, alcohol, and thiol provide
-formation of peptide bonds Classification: locations for a polar water molecule
Peptide bond 1. Whether the R group is acidic, basic, to hydrogen bond
-condensation reaction that is through neutral-polar, or neutral-nonpolar They are thus somewhat hydrophilic
dehydration synthesis that releases -AA with nonpolar R are classified as (like the OH groups in a sugar)
water and the new AA residue held hydrophobic These side chains are important in
together by a peptide bond -those with polar side chains are making a protein sufficiently water
-partially double bond character of classified as hydrophilic soluble to operate effectively inside a
the peptide bond 2. According to Timberlake cell
The O, C, N and H atoms are nearly -acidic polar: side chain contains a
planar and there is no rotation about carboxylic acid
the peptide bond
2
Acidic Polar
-Aspartic Acid
-Glutamic Acid
acidic AA have side chains that
contain a carboxyl group in addition
to the one next to the amino group
at cellular pH is near neutral (ph-7)
the carboxyl group is dissociated so
that the R group has a negative
charge (present as the carboxylate
anion)
secondary carboxylic group is weaker
acid
carboxylate side chain is important in
interaction with metal in many
enzymes, in ionic interactions
Basic Polar
-Lysine
-Arginine
-Histidine
side chain include an amino group.
These amino groups are also ionized
(present as the ammonium ion) at
neutral pH
Basic AA are positively charged as a
result of the dissociation of the amino
group in their side chain
The ionized groups are quite polar.
They make side chain quite
hydrophilic.
Acid and base chains are ionic and
therefore hydrophilic.
Lysine: ammonium ion
Arginine: guanidium group
Histidine: imidazolium group
Cystine formed by mild oxidation of
2 sulfihydryl group forming disulfide
bond
-hair perm creates disulfide linkages
Structure (Devlin)
Monoamino, monocarboxylic Gly, L
Ala
Unsubstituted: L-val, lue, ile
Heterocyclic: -Pro, -Phe
Aromatic: Tyr, Trp
Thiother: Met
Hydroxy: ser, thr
Mercapto: cys
Carboxamide: asn, L-gln
monoamino, dicarboxylic: L-asp, L-glu
diamino, monocarboxylic: L-lys, L-arg,
L-his
All 3 basic AA (KRH) have a positive
charge on the Nitrogen in the R side
chain
Serine is formed by adding a
hydroxyl group to Alanine
Cysteine is formed by replacing the
O with S in Serine
Threonine is formed by adding CH3
to Serine
Valine has a V shaped side chain
Leucine has a Y shaped side chain
Isoleucine has an upside down L-
shaped side chain
Proline is shaped like a pentagon
with the amino group incorporated in
the ring
cyclic amino acid. It is nonpolar (wt
aliphatic grp)
one of the ambivalent amino acids,
meaning that it can be inside or
outside of a protein molecule
occurs in proteins frequently in turns
or bends, which are often on the
surface
Gly: sharp bends, allow more
flexibility
When we replace the H with a methyl
group, we get Alanine
we add a phenyl group to alanine, we
get phenylalanine.
add a hydroxyl group to Phe, we get
Tyrosine
Aspartic acid is formed by adding a
carboxyl ion to Alanine:
Glutamic acid is formed by inserting
another CH2 into Aspartic acid:
Aromatic
nonpolar
To different degrees, all aromatic
amino acids absorb ultraviolet light.
Tyrosine and tryptophan absorb
more than do phenylalanine
tryptophan is responsible for most
of the absorbance of ultraviolet light (
280 nm) by proteins
Tyrosine is the only one of the
aromatic amino acids with an
ionizable side chain. Tyrosine is
one of three hydroxyl containing
amino acids
hydroxyl groups
serine and threonine are so high that
they are generally regarded as
nonionizing
Structure of Side Chain (Koolman)
1. Aliphatic (do not contain N, O, S in
side chain)
branched-chain AA or BCAA
sometimes used to refer to AA having
aliphatic side chains that are non-
linear
-Gly, -Ala, -Val, -Leu, -Ile
2. Sulfur containing
-Cys, -Met
3. Aromatic (benzene ring in side
chain)
-Phe, -Tyr, -Trp
4. Neutral (hydroxyl or amide groups in
side chain)
-Ser, -Thr, -Asn, -Gln
5. Acidic (carboxylate groups in side
chain)
-Asp, -Glu
6. Basic
-Lys, -Arg
7. Imino acid
-Pro
polar and positively charged at pH
values below their pKa's, and are very
hydrophilic
Lysine:
the side chain of lysine has a marked
hydrocarbon character, so it is often
found NEAR the surface, with the
amino group of the side chain in
contact with solvent
Histidine,
essential aa, has as a positively
charged imidazole functional group.
unprotonated imidazole is
nucleophilic and can serve as a
general base
protonated form can serve as a
general acid
serve a role in stabilizing the folded
structures of proteins
Cyclic AA
-Imino acid
-the only proteinogenic AA whose
side group links to the -amino group
and thus the only containing
secondary amine at the position
Most plant proteins have insufficient
amounts of lysine and tryptophan;
thus strict vegetarians should ensure
that their diet contains sufficient
amounts of these 2 AA
Isoelectric AA fully ionized and
internally neutralized by their own
amino and carboxyl groups
depending on the number of the
protons they can give up, we define
monoprotic, diprotic and triprotic acids
monoprotic, (e.g. acetic acid or
ammonium) have only one dissociable
group
diprotic (carbonic acid, bicarbonate,
glycine) have two dissociable groups
and
triprotic (e.g. phosphoric acid) have
three dissociable groups.
In the case of multiple pK values they
are designated by indices: pK1, pK2,
pK3, etc
For amino acids, the pK1 constant refers
to its carboxyl (-COOH) group
pK2 refers to its amino (-NH3) group
the pK3 is the pK value of its side chain
Both the a-amino group and the a-
carboxyl group are ionizable
a-COOH group: a weak acid, can
DONATE its proton, with a pKa of about
2-3
a-NH2 group: a weak base (there is an
unshared pair of electrons on the N; the
neutral amino group) can ACCEPT a
proton
side chain ionizable groups: only 7
amino acids
Asp, glu
His, lys, arg
Cys, tyr
3
 Glycine (Gly, G) the simplest and
smallest of all AA and the only one
which is not optically active since it
has a single H atom on its side chain
Alanine (Ala; A) has a methyl group
as its side chain.
Valine (Val; V) has a slightly longer
side chain again and this time there is
a branch. AS the aliphatic side chains
get longer they are also more
hydrophobic.
Leucine (Leu; L) is very similar to
Valine except it has another methyl
group attached to the side chain
Aromatic AA, aromatic ring as part of
their side chains, are highly hydrophobic.
Phenylalanine (Phe; F) first of all
the aromatic AA
-contains a phenyl ring attached to a
methylene group
Tyrosine (Tyr, Y) contains a hydroxyl
group at the end of the phenyl ring
(makes tyrosine less hydrophobic)
-is a reactive group (the side chains
so far have all been unreactive)
Sulfur containing
Cysteine (Cys; C) sulphydryl group
(-SH) is extremely reactive, can form
hydrogen bonds and disulphide
bridges
-even though the SH group can
form H bonds the long aliphatic part
of the side chain makes it quite
hydrophobic
Methionine (Met; M) is a very
special amino acid
-the start AA in the process of
translation therefore begins every
single protein
-Sulfur atom in a thioether linkage an
is relatively unreactive
-has a high hydrophobic side chain
Hydrophilic AA, neutral, those which are
acidic and those which are basic
Basic AA contain side chains which are
positively charged at physiological pH
-the pKa of Histidine which is between 6
and 7 in proteins means that it is able to
accept or donate proteins at physiological
pH. For this reason histidine is often found
at the active site of enzymes.
Lysine (Lys; K) has one of the
longest side chain
-although side chain appears to be a
hydrophobic hydrocarbon chain it is
very polar because of the terminal
amino group
Histidine (His; H) imidazole ring
which often sits inside the active site
of an enzyme (helps bond to be
broken or made)
-the pKa of Histidine is between 6
and 7 in proteins (is able to accept or
donate proteins at physiological pH)
-It can do this because it can exist in
2 states, uncharged or positively
charged
Arginine (Arg; R) has largest of all
side chains
-guanidino group attached to the side
chain
-has a high pKa value and is
positively charged at physiological pH
Neutral Polar AA,not charged at
physiological pH however they all have
groups in their side chains which are polar
and can form H bonds so are classed as
hydrophilic
Serine (Ser; S) contains an aliphatic
chain with a hydroxyl group
-the OH group makes the AA highly
reactive and hydrophilic as it readily
forms H bonds
Threonine (Thr; T) neutral AA which
has a highly reactive (and highly
hydrophilic) hydroxyl group
-contains 2 centers of asymmetry (2
asymmetric C atoms shared only by
isoleucine)
Asparagine (Asn; N) is the amide
derivative of Aspartic acid, side chain
is amidated
-resulting amide is uncharged
-there is a terminal amide group as
opposed to the carboxyl group on
aspartate
Glutamine (Gln; Q) is similar to Essential AA
Asparagine with a terminal amide 1. Histidine
(instead of a carboxyl group as in 2. Isoleucine
glutamate) 3. Leucine
-These 2 are called the amide 4. Lysine
derivatives of their parent AA 5. Methionine
6. Phenylalanine
Aromatic AA (Trp, Tyr, Phe) absorb 7. Threonine
light in the near UV region of the spectrum 8. Tryptophan
(250-300 mm) 9. Valine
Trp has highest molar absorptivity 10. Arginine (required for the young
followed by Tyr, with Phe making but not for adult)
only a small contribution
Disulfide bonds (between Cys Isoelectric point (pI)
residues in proteins) also absorb in pI = the pH exactly halfway between
the UV range, but much less than the the two pKa values surrounding the
aromatics zero net charge equivalence point on
the titration curve
Posttranslational modifications of AA
side chains If pH < pI, net charge is positive
Chemical modifications AFTER (more + than - charges)
biosynthesis of proteins If pH > pI, net charge is negative
Occur a few AA residues in some (more - than + charges)
proteins
AA
20 (biologically essential)
Can synthesize 12
-amphibolic intermediates of
glycolysis and CAC (9)
-3 (cysteine, tyrosine, hydrolysine)
from nutrionally essential
AA that we produce:
1. Alanine
2. Asparagine
3. Aspartic acid
4. Cysteine (from methionine)
5. Glutamic acid
6. Glutamine
7. Glycine
8. Proline
9. Serine
10. Tyrosine (Tyrosine is produced pI = pK1 + pK2/ 2
from phenylalanine, so if the diet = 2.34 + 9.60/2
is deficient in phenylalanine, = 5.97
tyrosine will be required)
At pH < pK1 COOH NH3 + +1
11. Arginine
12. Formed posttranslational At pH betw pK1 and pI half of COOH and
processing of collagen half COO and NH3 (0.5- and 1+)
-Hydroxylysine At pH betw pK1 and pK2 = COO, NH3+
-hydroxytyrosine 0
Above pK2 COO- and NH2 -1
4
 pH betw pK1 and pK2 COO, COOH,
NH3 : 0
pH >pK2 COO, COO,NH3 : -1
pH above pK3 COO, COO, NH2 : -2
pI
Dipolar form in which amino and
carboxyl are ionzed
Net charge = 0
The pH at which an AA is electrically
neutral : the sum of the + charges =
the sum of the neg charge
Region of buffereing : pKa +/- 1
Aspartic acid
COOH,NH3,COOH
At pH<pK1 COOH, COOH,NH3 : +1 ACIDS AND BASES
At pH bw pK1 &pK2 COO,COOH,NH3: 0 Arrhenius:
pH >pK2 COO, COO, NH3: -1 -that acids are compounds that
pH above pK3 COO, COO, NH2: -2 contain H and can dissolve in water to
relase H into solution
pK1 2.1 (+1 and 0)
-bases releases hydroxide ions (OH)
pK2 3.9 (0 and -1)
in solutuion
pK3 9.8 (-1 and -2) Bronsted-Lowry:
-Base: a molecule or ion that
pI = pK1 + pK2/2 ACCEPTS H ions from solution
= 2.1 + 3.9 -Acid: substances that can donate a H
= 3 ion
Lewis Model
-acid: accepts a pair of electrons
-base: donates a pair of electrons
Strong acids ionize completely in
water
Ex. Hydrochloric acid (HCl)
Weak acids ionize partially in water
Ex. Acetic acid (most of the acetic
acid in solution remains unionized and
a small fraction ionizes to form
CH3COO- and H3O+ ions
Strong bases ionize completely in
water.
Ex. Sodium hydroxide, NaOH
(contains entirely Na and OH ions)
pKa1 of Glutamic Acid = 2.2 (COOH)
pKa2 of Glutamic Acid = 4.3 (COOH) Weak Bases ionize partially in water
pKa3 of Glutamic Acid = 9.7 (NH2) -do not furnish OH-ions by
dissociation. They react with water to
pI=pK1 +pK2
furnish the OH ions
= 3.2 -when a weak base reacts with water
At pH <pK1 COOH,COOH,NH3: + the OH- comes from the water and
the remaining H+ attaches itself to
the weak base, giving weak acids as
one of the products
Ex. Ammonia, NH3+ (most of the
ammonia in solution remains
unionized and a small fraction ionizes
to form NH4+ and OH- ions
-when an acid reacts with a base
produces a salt and water, it is called
neutralization
Acid+Base Salt+water
pH and pOH are quantitative ways to
express solutions of acids and bases
Under the Bronsted-Lowry definition
both acids and bases are related to
the concentration of H ions
present
Acids increase the concentrations of H
ions
Bases decrease the concentration of
H ions (by accepting them
The acidity and basicity of something
therefore can be measured by its H
ion concentration
Concentration of H ion:
-measure of active acidity or basicity
-# of moles of H ion/liter of solution
-expressed as pH (the neg log to the
base 10 of H ion concentration)
pH scale is a more convenient way
than using exponential notation for
expressing concentration of H ions
-Danish biochemist Soren Sorensen
invented the pH scale for measuring
acifity. The pH scale is described by
the formula: pH=-log[H+]
-H ion concentration in units of moles
of H+ per liter of solution
For any Hydrogen ion concentration
of
- 1 x 10-n=[H+]
- the pH is equal to pH=n
-pH scale
The pH scale ranges from 0 to 14
pH and [H+] are inversely related-
lower means higher [H+]
therefore:
-substances with a pH between 0 and
less than 7: ACIDIC
-substances with a pH greater than 7
and up to 14 are BASES (higher pH
means lower [H+]
-right in the middle at pH 7 are
NEUTRAL substances, ex. Pure water
-NEUTRAL: H=OH (at 7pH under T 25
c)
pH affects functions, interactions
-change in pH may change
interactions
-dipole-dipole interactions might
change to ionic
-if pH of solution matches pKa (of
side chain) half is protonated and half
is deprotonated,
-at lower pH, more than half is
protonated
-at higher pH more than half is
deprotonated
Titration: methodof (usually) finding
the concentration of an unknown
liquid by comparing it with a known
liquid
Acid dissociation constant, Ka,
(acidity constant, or acid-
ionization constant)
-is a quantitative measure of the
strength of an acid in solution
-amount of H released
-specific type of equilibrium constant
that measures the propensity of
something larger to separate
(dissociate) reversibly into smaller
components
Ex.
HA A- + H+
-Ha represent acetic acid, and A- the
acetate ion
-the chemical species HA, A- and H+
are said to be in equilibrium when
their concentration do not change
with the passing of time
Equilibrium constant for the acid-base
equilibrium of an acid with its
conjugate base
Quotient of the qequilibrium
concentrations, denoted by [HA],[A-]
and [H+]
[ +][A]
Ka =
[]
When protonated and conjugated are
equal the conc of H is= to Ka
Conjugate base=undissociated
COO- = COOH
5
 K = H+ (conc.)
-logK = - log H
pK = pH
Prevailing H is numerically = to K
Experimental determination of pKa
values is commonly performed by
means of titrations
-calculate pH of weak acid after
addition of strong base (Henderson-
Hasselbach eq)
-quantitative relationship between pH
and conc weak acid and its base
[]
pH = pKa log
[]
Isoelectric AA fully ionized and
internally neutralized by their own
amino and carboxyl groups
Zwitterion: overall change is 0
Both the a-amino group (amino group
substituent on the aC) and the a-
carboxyl group (carboxyl substituent
on the aC) are ionizable
A-COOH group: a weak acid, can
DONATE its proton with a PKA of
about 2-3
A-NH2 group: weak base (theres an
unshared pair of electrons on the N:
the neutral amino group can ACCEPT
a proton)
Isoelectric point- pH at which the
predominant from zwitterion
PROTEINS
linear polymers built from 20
different L--amino acids
Peptide bond
Primary linkage of all protein
structures
chemical bond formed between the
carboxyl groups and amino groups
of neighboring amino acids
dehydration synthesis reaction
(condensation reaction)
The resulting CO-NH bond is called a
peptide bond, and the resulting
molecule is an amide (residue)
The four-atom functional group -
C(=O)NH- is called an amide group or
(in the context of proteins) a peptide
group
These bonds are rigid and planar, due
to electron sharing between the
carboxyl carbon and the amide
nitrogens which contribute to the bond
and give it a partial double-bond
character
Peptide bond is planar/flat: due to its
partial double-bond
The electrons from the double bond to
Oxygen migrate to the bond between th
C and the N producing partial + (N) and
-ve (O) charges
there is not free rotation about the C --
N bond
the group of atoms about the peptide
bond can exist in the cis or trans
configurations
The most stable conformation is planar
and trans
Proline is mostly in cis
Isomerization :about the C-N bond
Left : cis
Right: trans
chain torsion angles of a polypeptide
These are phi, psi, and omega (W)
Phi and psi (N-Ca and Ca-C bonds)
relatively are free to rotate
Free rotation:
Alpha C CO: Psi angle
Alpha C N: phi angle
Rachamandran angles
Free rotation can occur about the bonds
that connect the -C with the -N and with
the -carbonyl C
polypeptide chain is thus a semirigid
structure with two-thirds of the atoms
of the backbone held in a fixed planar
relationship
Each peptide chain has one amino
terminus and one carboxyl terminus.
These are the only free alpha amino and
alpha carboxyl groups in the peptide
All of the others are involved in the
peptide bonds between amino acids
peptide is a compound consisting of 2
or more amino acids
Oligopeptides have 10 or more amino
acids (20)
Polypeptides are chains of about
more 20 to 50
proteins are peptides consisting of
more than 100 amino acids
Classification accdg to composition
Simple
Consist solely of AA
Albumin
Globulin
Glutelin (glutenin-wheat, oryzenin-
rice
Protamine
Albuminoids, scleroprotiens: keratin,
elastin, collagen
Histones, globin
Conjugated
With non protein prosthetic grp
Nucleoprotein (nucleic acid with
protamine/histones) in chromatin
Glycoprotein/mucoprotein 9ground
substance of connective tissues)
Phosphoproteins ( H3PO4)
Chromoprotein( with hematin)
Hemoglobin, cytochromes,
rhodopsin
Lipoproteins
With Fats, phospholipids
Metalloproteins
Ceruloplasmin, siderophilin
Derived
Primary derivatives
Intramolecular rearrangement thru
hydrolysis
Denatured protein
Proteans
From action of water, dilute
acids or enzymes
Myosan, edestan
Metaproteans
From further nydrolysis
Acid albuminates/alkali
Coagulated
From action of heat,
alcohol, uv, shaking
Cooked egg albumin, meat
Secondary
From more extensive hydrolysis,
fragments of original ptotein which
are soluble in water, non
coagulable by heat
Primary proteoses
Peptones
peptides
Classification accdg to function
1. Structural proteins
Supporting filaments
Internal network : cytoskeleton,
cellular shape and physical integrity
Actin and myosin : contractile
machinery of muscles
examples
Collagen
elastin
Alpha keratin (hair, nail, feather)
Fibroin (silk of spider web)
Glycoprotein (cell coat)
mucoprotein

6
2. Transport solubility Interconnect 2 parallel chains thru Levels of Orders
Serum albumin Depends on the AA components on cysteine residues within each Primary structure:
Hemoglobin the surface polypeptide the amino acid sequence
Lipoprotein High hydrophobic on the surface : Relatively stable Aminoacyl residues: replace the suffixes
Ceruloplasmin low solubility with yl
Iron binding globulin Charged, polar surface residues Weak bonds Peptides are named as derivatives of the
3. enzymes - >2000 increase solubility 1. Hydrogen bonds carboxyl terminal aminoacyl residue
4. Nutrient and storage Sharing of bonds between the Lysyl-leucyl-glutamine
Ovalbumin Precipitation: salting out Nitrogen and carbonyl Oxygen of most fundamental, all proteins have
Ferritin Addition of neutral salt (NH4SO4) the same or different chains Amino acids that are ionizable in protein
Gliadin Increased reaction of charges of They are significant due to or polypeptide:
zein protein with the salt and exposes extremely large number of H bonds In side chain (7)
5. Contractile or motile hydrophobic areas on prtein Broken during denaturation - Basic: Histidine, arginine, lysine
Actin and myosin surface which aggregate and Hydrophobic bonds - Acidic: Aspartate, glutamate
6. Protective precipitate Nonpolar side chains of neutral - Thiol: Cysteine
Ig, fibrinogen, thrombin amino acids are associated with - tyrosine
7. Hormones Coagulation point one another Terminal amino acids
8. Toxins Characteristic of individual proteins Maintain protein structure
9. Receptors Acids lower Electrostatic bonds Secondary structure:
Alkali raises Salt bonds formed between Due to formation of H bonds between
Classification accdg to shape Hydration lowers oppositely charged groups in the one peptide bond with another within
1. globular Color reactions side chain of amino acids chain
Chains are tightly folded spherical Biuret : Protein in serum forms a Broken by denaturation carbonyl group (O) of amino-acid makes
molecules violet colored complex with cupric a hydrogen bond with the amino group
Most have motile function ions in an alkaline solution. The Folding (H) of another amino-acid
Soluble in aqueous solution intensity is proportional to the These linear chain of amino acids Dictated by the primary structure
Nearly all are enzymes amount of protein present interact with each other and their The most common examples are the
Antibodies Ampholytes surroundings in the cell to produce a alpha helix and
Nutrient storage In acid solution acts as base well-defined, three dimensional shape beta sheet
Hgb In alkaline acts as acid The shape into which a protein naturally Alpha helix
2. Fibrous folds is known as its native state R-handed coiled or spiral
Chains are arranged in parallel along Biological Roles of proteins The three-dimensional structure is conformation (helix), in which
a single axis to form long fibers or Acts as Catalysts. determined by the sequence of the every backbone N-H group donates
sheets Fibrous proteins serve as components of amino acids. a H bond to the backbone C=O
Physically tough the tissues holding the skeletal elements mechanism not understood group of the aa 4 residues apart
Insoluble in water together the mechanism depends equally on formed due to H bonds formed
Basic structural elements in The nucleoproteins serve as carriers of the characteristics of the cytosol, the between O of the CO and the H
connective tissues genetic characters and govern nature of the primary solvent (water atom of the peptide bond N of the
Collagen of tendon and bone inheritance. or lipid),
4th residue down the chain
matrix Performs transport function via catalytic macromolecular crowding
Alpha keratin of hair, skin, nails activity or as adsorbent. the concentration of salts backbone chain i is a R-handed
and feathers Acts as hormones and regulate the temperature helical conformation
Elastin of elastic tissues growth of plants and animals besides molecular chaperones assist in the Each turn of the helix comprises
3. those which fall between fibrous and controlling many other physiological folding of proteins 3.6 amino acids
globular functions. Most folded proteins have a hydrophobic stiff, rod-like structures
Long rodlike structures: myosin, Under condition of non-digestion the core in which side chain packing
fibrinogen protein accumulate inside cells and stabilizes the folded state
produce toxicity (e.g.Venoms of snakes the N-terminus of the protein begins to
Physical properties and insects). fold while the C-terminal portion of the
Tasteless, bitter (hydrolysates) protein is still being synthesized by the
Colorless, amorphous (some colored Bonds in the structure ribosome
and crystalline) Strong bonds
Insoluble in fat solvents 1. peptide
Varied solubility in water 2. Disulfide
amphoteric
7
 Proline induces bends in alpha helices
Since peptide bond N of proline lacks a
H atom to contribute to H bond, proline
can only be accomodated within first
turn of alpha helix
When present somewhere else it
disrupts the conformation of the helix,
producing a bend
Other AA that may disrupt regular
pattern of helix
Group of isoleucine causes steric
hindraance due to bulky side chains
Glycine due to small R which allows
movement destabilize
Group of acidic AA destabilize due to
negative charges which repel each
other
Lysine, Arg. Serine, threonine
Many alpha helices have predominantly
hydrophobic R on one side and
predominantly hydrophilic on the other
therefore amphipathic
Well adapted to formation of
interfaces between polar and
nonpolar regions
- Hydrophobic interior of a protein
and its aqueous environment
- Clusters can create channel, pore
that allow polar molecules to pass
thru hydrophobic cell membranes
Other helices
- the 310 helix and n helix
- ends of helices due to
unfavorable backbone packing in
the center of the helix
beta strand
(also strand) is a stretch of
polypeptide chain typically 3 to 10
aminoacids long with backbone in an
almost fully extended conformation
sheet refers to an assembly of at least
two such strands that are hydrogen-
bonded (or H-bonded) to each other
- between carbonyl O and amide H of
peptide bonds formed with adjacent
segments
Most are not perfectly flat but with
right-handed twist
Clusters of twisted strands form the core
of many globular proteins
in structure of the enzyme triose
phosphate isomerase
loops and turns are required to connect
alpha-helices and beta-strands
These are usually found on the surface
of the protein
Therefore they contain mainly
hydrophilic residues and are often
binding sites
Turns and bends:
Short segments of AA that join two units
of secondary structure (2 adjacent
strands of an antiparallel sheets)
A turn reverses the direction of a
helix/strand
is stabilised by a hydrogen bond
between the backbone CO of one
amino acid with the NH of the next
Beta turn: involves 4 residues in
which the 1st is H-bonded to the 4th
(tight 180-degree turn).
- Proline and glycine
A loop is a much larger sequence which
changes the direction of the helix/strand
There is no regular structure or
sequence of amino acids in these
regions
connect adjacent regions of
secondary structure
Irregular conformation
Tight turns and loose, flexible loops link
the more "regular" secondary structure
elements
Proline and glycine are known as "helix
breakers" because they disrupt the
regularity of the helical backbone
conformation
- have unusual conformational abilities
- commonly found in turns
Amino acids that prefer to adopt helical
conformations in proteins : methionine,
alanine, leucine, glutamate and lysine
prefer to adopt -strand conformations
- the large aromatic residues
(tryptophan, tyrosine and
phenylalanine)
- C-branched amino acids (isoleucine,
valine, and threonine)
supersecondary structure
formed by the combi of several
neighbouring alpha-helices/ beta-sheets
into a particular geometric arrangement
arrangements of 2 or 3 secondary
structures present in protein structures
term "motif" is often used to describe
super-secondary structures
connections between
long coiled-coil regions
alpha-helices connecting the
beta-sheets
refers to the overall three-dimensional
structure of a single polypeptide chain.
Regions of regular secondary structure
(e.g. alpha helicies and beta sheets) "
Tertiary structure
fold up" along with the "randomly"
coiled regions into a compact, generally
globular structure.
Tertiary structure is the "global" folding
of a single polypeptide chain
- to assemble the diff secondary struc
elements in a particular arrangement
- the overall shape of a single protein
molecule; often used as synonymous
with the term fold
- the spatial relationship of the
secondary structures to one another
domain is the unit of tertiary structure
Bonds that stabilize tertiary level
Hydrophobic
Ionic
H bonds
disulffide
controls the basic function of the protein
Quaternary structure
involves the association of two or more
polypeptide chains (tertiary structure)
into a (multi-subunit) active structure
Not all proteins exhibit quaternary
structure
Stabilized by;
D bonding, van der Walls interactions
ionic bonding (between charged Rs)
Hydrophobic (between non-polar Rs)
disulfide bonds between cysteine
residues in different polypeptide
chains
units of tertiary structure aggregate to
form homo- or hetero- multimers
Hetero-multimers
different tertiary domains aggregating
together to form a unit
Homo-multimers
more common to find copies of the
same tertiary domain associating
non-covalently
Separating proteins
Size
- Ultrafiltration
- centrifugation
- Size exclusion chromatography
through a chromatography
column filled with porous beads
Diff. Molecules enter the pores
according to how easily they can
enter
Larger pass thru column
Diff in the time required to pass
or the condition required to elute
the protein from the column
charge
- solubility, ion exchange
chromatography, and electrophoresis
- Proteins are least soluble at their
isoelectric point. At the isoelectric
point, many proteins precipitate from
solution
- Net charge (abov and below Pi)
ion exchange chromatography
and electrophoresis
- In ion exchange chromatography,
the greater the magnitude of the
charge, the slower a protein moves
through a column
- Electrophoresis
In electrostatic field
Molecules with no net charge do
not move
with a net positive charge move
toward the negative end
those with a net negative charge
move toward the positive end
magnitude of the net charge
determines how fast the species
moves
8
 Sequence of AA
- Denaturing the protein
cleaving the disulfide linkages (oxidation
of the S to SO3
Detrmine N terminal (Edman
degradation. tags the N-terminal residue
then cleaved)
-use Sangers reagent, dansyl chloride,
and leucine aminopeptidase
C terminal
- Akabori reaction (hydrazinolysis) and
reduction with lithium aluminum
hydride tag
- selectively cleave the C-terminal
residue using the enzyme
carboxypeptidase
cleave the polypeptide into smaller
fragments and determine the amino acid
composition and sequence of each
fragment
Partial acid hydrolysis randomly cleaves
the protein chain into a number of
fragments
Trypsin, a digestive enzyme, specifically
cleaves on the C-side of arginine or
lysine
chymotrypsin preferentially cleaves
residues containing aromatic rings
(tyrosine, phenylalanine, and
tryptophan). It slowly cleaves other
residues especially leucine. Clostripain
cleaves positively charged amino acids,
especially arginine; It cleaves lysine
more slowly
Chemical :
cyanogen bromide,
hydroxylamine, and heating an
acidic solution
Cyanogen bromide specifically
attacks methionine
Apply Edman degradation to the
fragments
Collagen
Main protein in CT of animals
Most abundant protein in mammals (25-
35%)
Tropocollagen: fundamental unit of
collagen (3 chains)
Alpha helix : L-handed helix
3 entwine to form R-handed triple helix
-Resists unwinding: it and its 3 chains
are coiled in opposite directions
located in the extracellular matrix of CT.
Most abundant protein in body
classification of an extracellular matrix
protein as a collagen is based on the
presence a distinctive triple-helical
conformation
The collagen triple helix consists of
three polypeptide chains super coiled
about a common axis and linked by
hydrogen bonds.
28 types
90% are type I-IV
Specific types are associated with
particular tissues
The most prevalent and well-studied
collagens belong to the fibril-forming or
interstitial collagen family
Type I
is the most common fibril-forming
collagen (90%)
Its fibrils make up the mineralized
matrix in bone, the strong parallel
bundles of fibers in tendon, and the
plywood-like alternating layers in the
transparent cornea
Dentin
fascia
Scar
skin
Type II
is the major fibril-forming collagen in
cartilage
Vitreous body, nucleus pulposus
Type III
reticulin
is found in blood vessels and skin,
together with type I, granulation tissue
Type IV
Basement membranes, which serve to
separate cell layers and act as filtration
barriers
- organized into a network or mesh-like
sheet structure
- In the kidney
Every 3rd residue is glycine ( gly-X-pro)
or Gly-X-Hyp) (X hydroxyproline or
hydroxylysine)
1/3 is glycine
Proline 9% of collagen
OH of Hyp : in forming H bonds
OH of Hyl attachment sites for
polysaccharides
Synthesis hemoglobin
In fibroblasts Metalloprotein (iron)
Translation of a-chains oxygen-transport in the red blood cells
(preprocollagen) (RER) of vertebrates
Hydroxylation (ER) rqrs vit C In mammals, the protein makes up
Glycosylation (ER) about 97% of the red blood cells dry
Exocytosis into extracellular content, and around 35% of the total
Outside fibroblast content (including water)
Cleavage of terminal of procollagen to four globular protein subunits
tropocollagen (insoluble) four polypeptide chains: two alpha
chains, each with 141 amino acids and
Tropocollagen chains associate to form two beta chains, each with 146 amino
microfibrils acids.
Cross linking covalent lysine- Each subunit is composed of a protein
hydroxylysine linkage chain tightly associated with a non-
protein heme group
Disorders
Ehlers Danlos syndrome Keratin
- Faulty collagen synthesis particularly abundant in the proteins of
Hyperextensible skin hair, hooves, and the keratin of the skin
Easy bruising
Hypermobile joints
Autosomial dominant or recessive
Type III most frequently affected
Osteogenesis imperfecta
Brittle bone disease
Autosomal dominant , most common
form
Abnormal type I ; (type II :fatal utero)
Multiple fractures (during birth)
Blue sclerae (translucent connective
tissue over choroid)
Hearing loss (abn middle ear bones)
Lack dentin dental abn
Alports sundrome
Abnormal type IV
(basement membrane of kidney, ears,
eyes)
Most common form X-linked recessive
Nephritis, deafness, ocular
Vit C deficiency
Prolyl and lysyl hydroxylases: poor
function, therefore no cross links
Myoglobin
a single-chain globular protein of 153
aa, containing a heme (Fe-containing
porphyrin) prosthetic grp in the center
around wc remaining apoprotein folds
8 alpha helices and a hydrophobic core
primary oxygen-carrying pigment of
muscle tissues.
9