Protein chemistry

512 :

.

1

Proteins
a

macromolecule that consists of one or

more polypeptide chains folded and coiled
into specific conformations
Made

up of various 20 amino acids

Vary widely in structure and function
Abundant about 50% of cellular dry weight

explain the biologically important

functions of PROTEINS
Structural support
2. Storage of amino acids
3. Transport (hemoglobin)
4. Signaling (chemical messengers)
5. Cellular response to chemical stimuli
(receptor proteins)
6. Movement (contractile proteins)
7. Defense against foreign substances &
disease-causing organisms (antibodies)
8. Catalysis of biochemical reactions (enzymes)
1.

biologically important
functions of PROTEINS

)
(
) (
)
(
) (
) -
(
) - (
) GH

4

What do proteins do?

off

on

Regulation

Movement

proteins

Signaling

Catalysis

Immune

Transport

Protein structure

1 Amino acids: structure, side chains

(charged, polar uncharged, nonpolar
aliphatic, aromatic)
2
Protein structure and function:
Structure: size and shapes, primary
secondary tertiary quaternary
(prosthetic groups) ; Biological functions
Structure & function: Domains, motif, and
family
3 Protein analysis
Purification Determine sequence, mass,
and structure (X-ray crystallography and
NMR)

1 Amino acids- basic structure

COOH3N

C
R

Common
structure
of 19 AAs

proline

-carbon is chiral (asymmetric) except in glycine (R is H)

2.

Amino acids are dipolar ions (zwitterions in aqueous solution and are
amphoteric

3.

The side chains (R) differ in size, shape, charge and chemical reactivity

4.

A few proteins contain nonstandard amino acids that are formed by posttranslational modification of proline and lysine.

10

1 Amino acids- charged (5)

Form salt bridges, are hydrophilic

1.Acidic amino acids (2): containing additional carboxyl groups which are
usually ionized

aspartic acid (Asp, D )

glutamic acid (Glu, E )

2. Basic amino acids (3): containing positively charged groups

Lysine (Lys, K)

Arginine (arg, R)
a guanidino group

Histidine (His, H )
The imidazole group has a pKa near neutrality.
This group can be reversibly protonated under
physiological conditions, which contribute to
the catalytic mechanism of many enzymes.

1 Amino acids- polar uncharged (5)

Contain groups that form hydrogen bonds with water,
hydrophilic

Serine (Ser, S)
Threonine (Thr, T)
Asparagine (Asn, N)
Glutamine (Gln, Q

Contain
hydroxyl
groups.

Cysteine (Cys, C

has a thiol group which is often oxidizes to

cystine

x-S-S-x

1 :Amino acids- nonpolar aliphatic (7)

(hydrophobic)

Glycine (Gly, G )

Proline (Pro, P

: imino acid

Methionine (Met, M )
: contains a sulfur atom

Alkyl side chains

Alanine (Ala, A)

Valine (Val, V)

Leucine (Leu, L )

Isoleucine (Ile, I )

1 Amino acids- aromatic (3)

Accounts for most of UV absorbance of proteins at 280 nm
hydrophobic

Phenylalanine (Phe, F )
Tyrosine (Tyr, Y)

Tryptophan (Trp, W)

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Peptides

19

Peptide
Peptide and peptide bond
A peptide bond is a covalent bond formed
between the carboxyl group of one AA and
the amino group of its next AA with the
elimination of one H2O molecule.

Peptides can be extended by adding

multiple AAs through multiple peptide
bonds in a sequential order.
dipeptide, tripeptide, oligopeptide, polypeptide

AAs in peptides are called as residues.

Learning Check
Write the name of the following tetrapeptide using amino acid
names and three-letter abbreviations.
CH3
CH3

H3N

CH CH3

SH

CH2

CH3 O

CH O

CH2 O

CH2 O

CH C N

CH C N

CH C N

CH C O

22

23

Learning Check
Draw the structural formula of each of the following peptides.
A.
Methionylaspartic acid
B.Alanyltryptophan
C. Methionylglutaminyllysine
D. Histidylglycylglutamylalanine

24

Biologically active
peptides
Peptides may act as hormones such as:
Vasopressin

and oxytocin hormones released by

pituitary gland.
Both have disulfide bridges.
Vasopressin (antidiuretic hormone) decreases urine
formation.
Oxytocin causes uterine contractions.

25

. Peptides may act as hormones such as :

Adrenocorticotropic

hormone released by

pituitary gland
Has no disulfide bridges
Regulates the production of steroids by the
adrenal gland

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Biologically active peptides

Glutathione (GSH):
L-glutamylcysteinylglycine
Glutathione is a tripeptide composed of
gamma glutamate, cystein, glycine.

Reduced glutathione (GSH)

maintains the normal reduced
state of the cell.

Glutathione: is a material found naturally in the human body

Peptides may act as reducing agents such as glutathione which is
needed for many enzymes. Glutathione is an abnormal tripeptide
(Glu-Cys-Gly) where Glu is linked to Cys by non a-peptide
bond.
-

NH3 +

O
O

H
N
O

N
H
SH

Glutathione, GSH
(reduced form)

O-

2e- oxidation
2e- reduction

NH3 +

H
N

S
S
N
H

N
H

H
N

NH3 +
O
Glutathione, GS-SG
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(oxidized form)

OO
O

A disulfide
bond
O-

Insulin
Insulin consists of
two polypeptide
chains,
A and B, held
together by two
disulfide bonds.
The A chain has
21 residues and
the B chain has 30
residues.
The sequence shown is
that of bovine insulin.
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 Glutathione

.
GSH

GSSG .
Coenzyme
Antioxidant
.Free radicals


.
30

H2O2

2GSH
GSH
peroxidase

2H2O

GSSG

NADP+
GSH can be
regenerated from
GSH reductase
GSSG by the enzyme
glutathione reductase
(GSR(

NADPH+H+

As a reductant to protect nucleic acids and

proteins from toxin by discharging free radical
or H2O2

2-Protein structure and

function

32

2-Protein structure and function

Structure: size and shapes,
primarysecondary tertiary
quaternary, prosthetic groups
nonprotein molecules of conjugated
proteins
Domains, motif, and family
Protein function

2 Protein structure Sizes

1. A few thousands Daltons (x 103) to more
than 5 million Daltons (x 106)
2. Some proteins contain bound nonprotein
materials (prosthetic groups or other
macromolecules), which accounts for the
increased sizes and functionalities of the
protein complexs.

2-Protein structure Shapes

Globular proteins: enzymes

Complementary fit of a substrate molecule to the catalytic site

on an enzyme molecule.

36

Fibrous proteins: important structural

proteins (silk fibroin, keratin in hair and
wools )

Protofibril

Keratin

keratin in hair
.

microfibril

38

39

2 Protein structure Primary

Polypeptides contain N- and C- termini and are
directional, usually ranging from 100-1500 aa

.
.
.
4 )
574

Formation of a peptide bond (shaded in gray) in a dipeptide.

N terminus
C terminus

Structure of the pentapeptide Ser-Gly-Tyr-Ala-Leu.

Secondary
structure -

:
-
) ( .
- )
(.
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-helix

right-handed
3.6 aa per turn
hydrogen bond
N-HO=C

A stereo, space-filling representation

Collagen triple helix:

three polypeptide intertwined

-sheet:

hydrogen bonding of the pepetide bond N-H and

C=O groups to the complementary groups of
another section of the polypeptide chain

Parallel sheet: sections run in the

same direction
Antiparallel sheet: sections run in
the opposite direction

Protein structure Tertiary

The different sections of -helix, -sheet, other minor secondary
structure and connecting loops of a polypeptide fold in three dimensions

.
.
. :
- . .
-
. .
- . .
- .

Noncovalent interaction between side chains that

hold the tertiary structure together: van der Waals
forces, hydrogen bonds, electrostatic salt
bridges, hydrophobic interactions
Covalent interaction: disulfide bonds

Interactions stabilizing proteins

Myoglobin (Mb)
Located in muscle to supply O2
1st protein in high resolution
153 AAs
75% of structure is -helix in 8
regions.
the interior almost entirely nonpolar
residues

Ribonuclease

A pancreatic enzyme that hydrolyzes RNA

124 AAs
Mainly -sheet
Highly compact and nonpolar interior
4 disulfide bonds

Rhodopsin
Photoreceptor
protein
7 transmembrane
helices
11-cis-retinal
chromophore
in the pocket
Residues are
modified.

Protein structure Quaternary

Many proteins are composed of two or more polypeptide chains (subunits).
These subunits may be identical or different. The same forces which stabilize tertiary
structure hold these subunits together. This level of organization called quaternary
structure.

. :

.

Levels of Protein Structure

Primary:

the sequence of the amino

acids in the chain and the disulfide links.
Secondary: structure formed by
hydrogen bonding. Examples are -helix
and pleated sheet.
Tertiary: complete 3-D conformation.
Quaternary: association of two or more
peptide chains to form protein.
51

The Four Levels of Protein Structure

Protein Structure and Function

52

Protein structure
Prosthetic groups
Covalently or noncovalently attached to
many conjugated proteins, and give the
proteins chemical functionality.
Many are co-factors in enzyme reactions.
Examples : heme groups in hemogobin

Protein structure
Motifs, Domains, and Families

Supersecondary structures
Supersecondary
structures = motifs
Combinations of 2
structures
Functional
Structural
Helix-turn-helix
Hairpin (Connects
antiparallel
Oftenstrands)
have functional significance and represent
the essential
Greek keyparts of binding or catalytic sites
conserved
-- among a protein family

55

 motif

beta-ribbon
the simple protein structural motif involving
two beta strands that look like a hairpin

Common motifs

Greek key

Protein domains
many globular proteins consist of several compact, locally folded and
stable regions called domains i.e. modular units.
The tertiary structure of many proteins is built from
several domains.
Often each domain has a separate function to perform
for the protein, such as:
binding a small ligand
factors)

* DNA-binding (in transcription

spanning the plasma membrane (transmembrane

proteins)

containing the catalytic site (enzymes)

providing a surface to bind specifically to another
protein

57

57

Protein Domains

Most proteins are composed of structural

subunits called domains
A domain is a compact unit of protein
structure, usually associated with a
function.
It is usually a fold - in the case of
monomeric soluble proteins.
A domain comprises normally only one
protein chain: rare examples involving 2
chains are known.
Domains can be shared between
different proteins: like a LEGO block

Protein families: structurally and functionally related proteins from different

sources

Motif

The primary structures of c-type cytochromes from different

organisms

Classification of proteins
proteins can be classified according to structure
Simple Proteins: yield only amino acids on
hydrolysis
Conjugated (Complex )Proteins contains other
substances in addition to A.A.
which are much more common than simple
proteins, yield other compounds such as
carbohydrates, fats, or nucleic acids in addition
to amino acids on hydrolysis
60

Classification of proteins

Complex or conjugated protein include:

Nucleoprotein: pr + nucleic acids (chromatin)
Phosphoprotein: pr + phosphate group
(casin of milk)
Glycoprotein: pr + CHO (antibodies, mucin,
blood group, TSH..)
Lipoprotein: pr + lipids (plasma lipoproteins)
Metalloprotein: pr + metal (insulin, ferritin,
ceruloplasmin)
Chromoprotein: pr + coloured substance
which is a protoporphyrin containing metal
(Hb, chlorophyl)
61
Protein
Structure and Function

Physical and Chemical

Properties of Proteins

Lets examine the behavior of a Weak Acid( HA), in aqueous solution.

Relates three terms: pH, pKa, and [A-]/[HA]. If you

know two of these values, you can determine
the third.
pH = pKa + log([A-]/[HA])
When [A-] = [HA]:
pH = pKa + log(1)
pH = pKa + 0
pH = pKa

pka is the pH at
which a functional
group exists 50% in
its protonated
form (HA) and 50%
in its
deprotonated

63

Amphoteric
Isoelectric point
AAs in solution at certain pH are predominantly in
dipolar form, fully ionized but without net
charge due to -COO- and -NH3+ groups.
This characteristic pH is called isoelectric point,
designated as pI.
pI is determined by pK, the ionization constant of
the ionizable groups.

R CH COOH
NH2

+OH
R CH COOH

NH3+

+H+

+OH
R CH COOR CH COO

NH3+

+H+

NH2

pH<pI

pH=pI

pH>pI

cation

amphoteric

anion

Amino acid

pI

M.W.

Glycine

5.97

75

Alanine

6.00

Valine

Amino acid

pI

M.W.

cystein

5.07

121

89

methionine

5.74

149

5.96

117

asparagine

5.41

132

Leucine

5.98

131

glutamine

5.65

146

Isoleucine

6.02

131

cystein

5.60

119

Phenylalani
ne

5.48

165

aspartic
acid

2.97

133

Proline

6.30

115

glutamic
acid

3.22

147

tryptophan

5.89

204

Lysine

9.74

146

serine

5.68

105

Arginine

10.76

174

tyrosine

5.66

181

Histidine

7.59

155

pI Depends on Side Chain

The 15 amino acids thiol, hydroxyl groups or pure hydrocarbon
side chains have pI = 5.0 to 6.5 (average of the pKas)
Glu and Asp have acidic side chains and a lower PI
His, Arg, Lys have basic side chains and higher pI

 Side-chains

of a protein have many

ionizable groups, making the protein
either positively or negatively
charged in response to the pH of the
solution.
The pH at which the protein has zero
net-charge is referred to as
isoelectric point (pI).

COO-

COOH
+ OH-

P
NH3+

+ OH-

+ H+

NH3

cation

amphoteric

pH < pI

pH = pI

COO-

+ H+

NH2
anion
pH > pI

pI of most protein is ~ 5.0, and negatively charges in

body fluid (pH7.4)
pI > 7.4: basic proteins: protamine, histone
pI < 7.4: acidic proteins: pepsin

Colloid property
Diameter:

1~100nm,
in the range of
colloid;

Hydrophilic

groups
on the surface form a
hydration shell;

Hydration

shell and
electric repulsion
make proteins stable
in solution.

- - + +
+ - +
- + - +
- +-+
+ - +
+
- +
-

Precipitation of protein colloid

+
+

+ +
+
+ +

positively charged
(hydrophilic)
dehydration

+ + +
+
+
+
+ +
positively charged
(hydrophobic)

acid

base

base

acid

isoelectric point
(hydrophilic)
dehydration

base

Instable protein
(deposition)

negatively charged
(hydrophilic)
dehydration

acid

negatively charged
(hydrophobic)

Protein denaturation
The process in which a protein loses its
native conformation under the treatment
of denaturants is referred to as protein
denaturation.



.

The denatured proteins tend to

- decrease in solubility;
- increase the viscosity;
- lose the biological activity;
- lose crystalizability;
- be susceptible to enzymatic digestion.

PROTEIN DENATURATION

What are some factors that can lead to protein denaturation?

Question
At pH 7, which of the following amino acids
have a net positive charge, which have a net
negative charge, and which are neutral?
Lysine
Phenylalanine
Leucine

How can denaturation be classified regarding its

reversibility?
Protein denaturation can be a reversible or an
irreversible process, i.e., it may be possible or
impossible to make the protein regain its original spatial
conformation

What is protein denaturation? Is there any change in

the primary structure when a protein is denatured?
Secondary, tertiary and quaternary structures of proteins
are spatial structures. Denaturation is modification in
any of these spatial structures that makes the protein
deficient or biologically inactive.After denaturation the
primary protein structure is not affected.

75

In sickle cell anemia, a hereditary disease, there is substitution of

one amino acid by another in one of the four polypeptide chains of
hemoglobin. In this case are all of the structural levels of the
protein modified?
In sickle cell disease there is a change in the primary protein structure of
one of the polypeptide chains that form hemoglobin: the amino acid
glutamic acid is substituted by the amino acid valine in the chain. The
spatial conformation of the molecule in addition is also affected and
modified by this primary mistake and the modification also creates a
different (sickle) shape to the red blood cells.
What is the difference between essential and natural amino acids?
Essential amino acids are those that the organism is not able to
synthesize and that need to be ingested by the individual. Natural amino
acids are those that are produced by the organism.
The essential amino acids for humans are phenylalanine, histidine,
isoleucine, lysine, methionine, threonine, tryptophane and valine.
76

- Explain How:
a molecule's charge (amino acids) changes with the
pH of its surroundings.

THANK YOU