Intended Learning Outcomes

• At the end of the discussion, the

student must be able:
▫ To understand the life cycle of protein;
▫ To identify the four level structure of
proteins;
▫ To define protein denaturation; and
▫ Clinical correlation of different types of
proteins.
Protein Cycle
FOUR LEVELS OF
PROTEIN STRUCTURE
• Amino acids can undergo
condensation reactions in
any order, thus making it
possible to form large
numbers of proteins.
• Structurally, proteins can
be described in four ways.
▫ Primary
▫ Secondary
▫ Tertiary
▫ Quaternary structure
FOUR 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 β-pleated sheet.
• Tertiary: complete 3-D conformation.
• Quaternary: association of two or more
peptide chains to form protein.
Protein Structures or CONFORMATIONS

Hydrogen bond

Pleated sheet
Polypeptide
Amino acid (single subunit)

(a) Primary structure

Hydrogen bond

Alpha helix

(b) (c) Tertiary

Secondary structure
structure

(d) Quaternary structure

Protein Hierarchy
 Primary Structure

The primary structure of a protein is its linear

sequence of amino acids and the location of any
disulfide (-S-S-) bridges.
 Primary Structure
• The primary structure of a protein is defined
by the sequence of amino acids, which form
the protein. This sequence is determined by
the base pair sequence in the DNA used to
create it.
Naming a Peptide Chain
• “residue” or “moiety” – each component amino
acid
• Amino acid name ending in –in, ine, ic, an, and
–ate
▫ Changed to –yl except the C-terminal
• Tri-, or octa- refers to the number of residues,
not the number of bonds.
 Home Activity No.1

• Make a peptide chain of the following:

▫ Glu-Ala-Lys-Gly-Tyr-Ala
▫ First 5 letters of your first name.
 Secondary Structure
• The secondary structure describes the way that
the chain of amino acids folds itself due to
intramolecular hydrogen bonding.
• Two common secondary structures are :
the a-Helix →

and the b- sheet →

Alpha Helix
Pleated Sheet
 Alpha Helix

• This orientation of H-bonding produces a helical coiling

of the peptide backbone such that the R-groups lie on the
exterior of the helix and perpendicular to its axis.
 Alpha Helix
• Amino acids such as A, D, E, I, L and M
favor the formation of -helices.
• G and P favor disruption of the helix
(producing a bend).
• The disruption of the helix is important as
it introduces additional folding of the
polypeptide backbone to allow the
formation of globular proteins.
 Top view of alpha helix
Angle: C-N bond
-57°
is termed phi()
angle

0.54 nm
(3.6 residues)

0.15 nm

Four carbonyl groups are pointing

upwards spaced roughly 100° apart on -47°
the circle, corresponding to 3.6 amino-
acid residues per turn of the helix. CO- C bond is
psi () angle
 Proteins in -helix
• KERATIN – fibrous protein whose structure is
nearly entirely -helical.
• HEMOGLOBIN – a globular, flexible molecule
whose structure is approximately 80% -helical.
 The Beta pleated sheet
• Composed of 2 or more
different regions of
stretches of at least 5-10
amino acids.
• Stabilized by H-bonding
between amide N’s and
carbonyl C’s.
• H-bonding residues are
present in adjacently
opposed stretches of the
polypeptide backbone.
Pleated -Sheets
Parallel and Anti-Parallel Sheets
Parallel and Anti-Parallel Sheets
• When the H-bonds are formed between
the polypeptide backbones of separate
polypeptide chain, they are termed
INTERCHAIN BONDS.
• The H-bonds of a -sheet formed by a
single polypeptide chain folding back on
itself are termed INTRACHAIN
BONDS.
-sheet PROTEIN
• Found in both fibrous
and globular proteins
• AMYLOID PROTEIN –
composed of twisted -
pleated sheet fibrils
whose 3D structure is
identical to that of silk
fibrils.
LOOPS, TURNS and BENDS

• Loops are regions that

contain residues beyond
the minimum number
necessary to connect
adjacent regions of
secondary structure.
two α helices (blue) connected
by a loop (red)
LOOPS, TURNS and BENDS
• TURNS and BENDS refer to short
segments of amino acids that join
two units of secondary structure
(e.g., 2 adjacent strands of
antiparallel -sheet).
•  turn – involves 4 aminoacyl
residues, in which the 1st residue is
H-bonded to the 4th, resulting in a
180° turn.
• Proline and Glycine are often
present in  turns.
Supersecondary Structures or Folds
• Also known as Structural motifs

Zinc finger

α helix and an antiparallel β

sheet zinc ion (green) is
coordinated by two histidine
Helix-loop-helix
residues and two cysteine
residues.
Supersecondary Structures or Folds

The λ repressor of bacteriophage

Leucine Zipper (blue) lambda employs a helix-turn-
bound to DNA. The leucine helix (green) to bind DNA (blue
residues are colored red and red)
FOUR MOST COMMON MOTIFS
anti-parallel -pleated sheet
FOUR MOST COMMON MOTIFS
 Tertiary Structure
• The tertiary structure
maintains the three
dimensional shape of the
protein (give shape to the
secondary structure).
• The amino acid chain (in
the helical, pleated or
random coil form) links
itself in places to form the
unique twisted or folded
shape of the protein.
 Tertiary Structure
• There are four ways in which parts of the amino
acid chains interact to stabilize its tertiary shape.
These includes:
• Covalent bonding: e.g disulfide bridges formed
when two cysteine molecules combine in which the –SH
groups are oxidized
• Hydrogen bonding between polar groups on the side
chain.
• Salt bridges (ionic bonds) formed between –NH3+
and –COO- groups
• Hydrophobic/Hydrophillic interactions
 Functions of Domain
• Binding of a substrate or other ligands.
• Anchor a protein to a membrane.
• Interact with a regulatory molecule that
modulates its function.

Triosephosphate
isomerase (beta barrel) HMG-CoA reductase
The three domains of pyruvate kinase
Proteins with a 3D structure
fall into two main types:
• Globular - These tend to form ball-like
structures where hydrophobic parts are towards
the center and hydrophilic are towards the
edges, which makes them water soluble.

• Fibrous - They proteins form long fibers and

mostly consist of repeated sequences of amino
acids which are insoluble in water.
FORCES CONTROLLING PROTEIN
STRUCTURES
 FORCES CONTROLLING PROTEIN
STRUCTURES
III. Electrostatic Forces
3. Dipole-dipole
Three types
1. Charge-charge
✓ Favor protein folding.
✓ Between oppositely
charged R-groups such
as K or R and D or E.
2. Charge-dipole
✓ Interaction of ionized R-
groups of amino acids
with the dipole of the
water molecule.
✓ The slight dipole
moment that exist in
the polar R-groups of
amino acid also
influences their
interaction with water.
✓ Majority of the amino
acids found on the
exterior surfaces of
globular proteins
contain charged or
polar R-groups.
 FORCES CONTROLLING PROTEIN
STRUCTURES

IV. van der waals Forces

▫ Attractive van der Waals forces involve the
interactions among induced dipoles that arise
from fluctuations in the charge densities that
occur between adjacent uncharged non-bonded
atoms.
▫ Repulsive van der Waals forces involve the
interactions that occur when uncharged non-
bonded atoms come very close together but do not
induce dipoles.
Disulfide (S-S) bonds
Quaternary Structure
 Quaternary Structure
• Many proteins are not single strands
• The diagram below shows the quaternary structure of
an enzyme having four interwoven amino acid strands.
 Quaternary Structure
• Oligomeric proteins - proteins with
multiple polypeptide chains; can be
composed of multiple identical
polypeptide chains or multiple distinct
polypeptide chains.
▫ Homooligomers – proteins with identical
subunits.
▫ Heterooligomers – proteins containing
several distinct polypeptide chains.
 HEMOGLOBIN
• The oxygen carrying protein of the blood.
• Contains two  and two  subunits arranged with a
quaternary structure in the form, 22
• A ___________ protein.
 PROTEIN FOLDING
• Protein fold and unfold in milliseconds;
• Unfolding and refolding occur 100 or 1000 times
during their lifetime;
• Ribosomes participate the first time the
protein is folded, but not in subsequent folding;
• Increase concentration of proteins affect the
kinetics of protein folding; and
• Information needed for correct protein folding
is contained in the primary structures.
 Factors that facilitate folding
and refolding
I. Native conformation of a protein is
thermodynamically favored
II. Folding is modular.
Stage 1 Stage 2
Newly synthesized Hydrophobic regions Native
polypeptide folds into are driven into the conformation
secondary structure interior of the protein
and away from the
solvent
Secondary structures
facilitates proper
folding
Factors that facilitate folding
and refolding
III. Auxilliary proteins assist folding:
a. Chaperone proteins
b. Protein disulfide isomerase
c. Proline - cis, trans - isomerase
 CHAPERONES
• Specialized group of
protein required for the
proper folding of many
species of proteins.
• PCB (Polypeptide
chain-binding) protein.
• Acts as catalysts by
increasing the rates of
the final stage in the
folding process.
 CHAPERONES
• Do not convey steric information.
• Do not form part of the final structure.
• Suppress non-productive interactions by binding
to transiently exposed portions of the
polypeptide chain.
• First identified as heat shock proteins (Hsp).
• Hsp expression is elevated when cells are grown
at higher-than-normal temperatures.
• Use an ATP-dependent mechanism.
Major Types of CHAPERONES
• Hsp70 (cytoplasm, ER,
chloroplasts, mitochondria):
▫ thought to bind and stabilize the
nascent polypeptide chain as it
is being extruded from the
ribosome.
▫ also involved in "pulling" newly
synthesized polypeptide into ER
lumen.
▫ bind short sequences of
hydrophobic amino acids thus,
shielding them from solvent.
Major Types of CHAPERONES

• Hsp60
(mitochondria,
chloroplasts)
▫ forms large 28-subunit
complexes called GroEL.

• Chaperonins: provides a sheltered

environment in which a polypeptide can fold
until all hydrophobic regions are buried in its
interior, thus eliminating aggregation.
 PROTEIN DISULFIDE
ISOMERASE
• Facilitates formation of disulfide bonds that
stabilize a protein’s native conformation.

• PROLINE – cis, trans – ISOMERASE

▫ Catalyzed isomerization from trans to cis
▫ Cis configuration is common in -turns
IV. Folding is a Dynamic Process
• Unfold refold
• Chaperone “rescue” unfold proteins.
• Glutathione reduces inappropriate
disulfide bond.
PROTEIN DENATURATION
• Disruption of the normal structure of a protein, such
that it loses biological activity.
• Some proteins will return to their native structures
under proper conditions; but extreme conditions,
such as strong heating, usually cause irreversible
change.
PROTEIN DENATURATION
Changes in temperature & pH can
denature (unfold) a protein so it no
longer works.
Cooking denatures
protein in eggs

Milk protein separates into

curds & whey when it
denatures.
PROTEIN DENATURATION
• Caused by ___________?
• EFFECT: unfolding and disorganization of
protein structure (not accompanied by
hydrolysis of peptide bond).
 PROTEIN DENATURATION
Agents Effect
Heat hydrogen bonds are broken by increased translational
and vibrational energy. (coagulation of egg white
albumin on frying.)
Ultraviolet Radiation Similar to heat
(sunburn)
Strong Acids or Bases salt formation; disruption of hydrogen bonds.
(skin blisters and burns, protein precipitation.)
Urea competition for hydrogen bonds.
(precipitation of soluble proteins.)
Some Organic (e.g. ethanol & acetone) change in dielectric constant and
Solvents hydration of ionic groups.
(disinfectant action and precipitation of protein.)
Agitation shearing of hydrogen bonds.
(beating egg white albumin into a meringue.)
PROTEIN DENATURATION
Reversible or Irreversible?
• REVERSIBLE
▫ In rare cases, upon removal of denaturing agents.
▫ Protein refolds into original native structure.
• PERMANENT
▫ Most proteins remain permanently disordered.

Denatured proteins are

often insoluble and
therefore precipitate
from solution.
Pathologic Consequences of Protein
Conformation Perturbation
• PRION DISEASES
▫ Transmissible spongiform encephalopathies.
▫ Fatal neurodegenarative diseases characterized by
spongiform changes, Astrocytic gliomas, and neuronal
loss resulting from the deposition of insoluble protein
aggregates and neural cell.
• Includes:
a) Creutzfeldt-Jakob disease (Human)
b) Scrapies (sheep)
c) Alzheimer's disease
d) Bovine spongiform encephalopathy (Mad Cow
Disease) in cattle
e) *Scurvy
PRION PrPSc (for scrapie)
PrPc (for cellular) • The abnormal, disease-
• The normal protein has its producing protein
secondary structure • Primary structures are
dominated by alpha helices identical but its secondary
(probably 3 of them) structure is dominated by beta
• is easily soluble conformation
• is easily digested by • is insoluble in all but the
proteases strongest solvents is highly
• is encoded by a gene resistant to digestion by
designated (in humans) proteases
PRNP located on our • When PrPSc comes in contact
chromosome 20. with PrPC, it converts the
PrPC into more of itself
• These molecules bind to each
other forming aggregates.
 ALZHEIMER’S DISEASE
• Refolding or misfolding of  - amyloid in human
brain tissue.
• Elevated levels of  - amyloid undergoes
conformational transformation.

Soluble -helix Apolipoprotein E - sheet rich state

rich state (prone to self
aggregation)
 BETA - THALASSEMIA
• Genetic defects that impair the synthesis
of one polypeptide sub-units of
hemoglobin.
Sickle Cell Anemia
• A small change in the
sequence of the primary
structure can have a
significant impact on protein
structure
• In sickle cell anemia a
glutamic acid is replaced by a
valine in the amino acid
sequence
 Changing Amino Acid Sequence
•Substitution of one amino acid for another in
hemoglobin causes sickle-cell disease.

2 7. . . 146
1 3 6
4 5
(a) Normal red blood cell Normal hemoglobin

2 7. . . 146
1 3 6
4 5
(b) Sickled red blood cell Sickle-cell hemoglobin
 Other Important Proteins
• Blood sugar level is
controlled by a protein called
_______.
• Insulin causes the liver to
uptake and store excess sugar
as glycogen.
• The cell membrane also
contains proteins.
• Receptor proteins help cells
recognize other cells.