Transcript Protein structure and Function

Protein structure and Function
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Objectives:
Structure of amino acids as a building
unit of proteins.
Acidic and basic properties of amino
acids.
Levels of protein structure
Protein misfolding.
Protein structure and Function
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Proteins are the most abundant and
functionally diverse molecules
1. function: enzymes, hormones, protein in
muscle, receptors, Hb & Mg
2. Structure: hair, nail, bone, skin…. etc
Amino acid (AA) structure:
AA are building blocks of proteins
Only 20 AA are found in mammalian proteins
& much more are found in nature (<300)
General Structure
of amino acid
Classification of AA
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AA with Nopolar side chains
AA with polar uncharged AA
Acidic AA
Basic AA
AA with imino group
You should know names, structures,
three letters & one letter
AA with nonpolar side chains
AA with nonpolar side chains
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Each of these AA has a non polar side chain that
does not give off proton or participate in hydrogen or
ionic bonds
The non polar R group fill up the anterior of the
folded protein and give it its three-dimensional
shape.
In proteins that are located in a hydrophobic
environment, such as membrane, the non-polar R
group are found on the outside surface of the
protein, interacting with the lipid content. This plays
an important role in stabilizing protein strucure.
AA with Uncharged Polar Side
Chains
AA with Uncharged Polar Side
Chains
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These AA have zero net charge at neutral pH.
Serine and threonine: Each contains a polar hydroxyl group
that can participate in hydrogen bond formation.
Additionally this polar hydroxyl group can serve as a site of
attachment of structure such as phosphate group or an
important component of active site of many enzymes.
Asparagine and glutamine: Each contain carbonyle group
and amide group can participate in hydrogen bond.
Moreover it can serve as asite of attachment of
oligosaccharide chains in glycoproteins. As well as serine
Tyrosine: has phenolic group that carries negative charge at
pH above its PKª ( pH = 10.5), so it is not hydrophobic at
this pH range.
AA with Uncharged Polar Side
Chains
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Cysteine: In proteins, SH group
(sulfhydry) of two cysteine become
oxidized to form dimer Cystine, which
contains covalent cross-link called
disulfide bond (S-S).
AA with acidic side chain
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They are proton donors.
At physiological pH, these AA are
negatively charged ( - ve are present on
the side chain.
-Aspartic acid
-Glutamic acid
AA with basic side chain
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They are proton acceptors.
At physiological pH, these AA are
positively charged ( + ve are present on
the side chain.
-Histidine
-Arginine
-Lysine
Proline
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Side chain of proline and its α -amino
group form a ring structure, so proline
differ from other amino acids. It
contains Imino group
Unique geometry of proline contributes
to the formation fibrous structure of
collagen and interrupts of the α -helix
found in globular proteins
Metabolic classification of amino
acids
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Ketogenic or glucogenic amino acids.
1- Ketogenic amino acids whose
catabolism yields either acetoacetate
or one of its precursors.
2- Glucogenic amino acids whose
catabolism yields pyruvate or one of
the intermediate of citric acid cycle.
Biological classification of Amino
Acids
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Essential or nonessential amino acids.
Optical Properties of AA
α -carbon of AA is attached to four
different groups & therefore is optically
active.
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Enantiomers or
Stereoisomers
 All AA are optically
active except Gly
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Optical Properties of AA
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Amino acids have asymetric center on
α-carbon atom so they exist in two
forms L & D forms.
All amino acids found in mammalian
proteins are L- form, however Dform is found in bacterial cells.
Optical Properties of AA
O
H2N
CH
C
OH
R
Proteins are made of amino acids
- a-amino acids
- only L-isomers found
Acidic and Basic Properties of
AA
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Acids: Compound that donate protons.
Bases: Compound that accept protons.
Strong acids: Ex, Hcl which dissociated
completely.
Weak acids: Ex, Acetic acid which dissociate
only to a limited extend.
Buffer: It is a solution that resists change in
pH following the addition of an acid or base.
It can creat by mixing a week acid and its
conjugate base.
Acidic and Basic Properties of
AA
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AA in aqueous solution contain weekly
acidic α-carboxyl & α-amino groups in
addition to acidic & basic side chains AA
Henderson-Hasselbalch equation:
HA
H+ + A -
Weak acid
conjugate base
Ka= [H+] [A-]
[HA]
Acidic and Basic Properties of AA
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Taking the logarithmic of both sides of
the equation.
Multiplying both sides of equation by -1
Substitution pH= - log [H+] , and
pKa = -log ka we obtain HendersonHasselbalch equation
Acidic and Basic Properties of AA
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pH = pKa + log [A-]
[HA]
The larger Ka & the smaller pKa ,the stronger the acid
& vice versa
Buffers: solution that resists change in pH following
addition of acid or base.
Maximum buffering capacity occurs at pH=pKa &
effective for ± 1 pH unit of pKa.
At pH values less than pKa, protonated acid form is
predominant. At pH values greater than pKa ,
deprotonated base form is predominant in solution.
Titration of Acetic Acid
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A solution containing acetic
acid (HA=CH3CooH),and
acetate(A- = CH3Coo-) with a
PKa of 3.8 resists a change
in pH from 3.8 to 5.8 with
maximum buffering capacity
at pH 4.8.
At pH< PKa the protenated
acid form is predominant
(CH3CooH)
At pH> PKa the
deprotenated base form is
predominant (CH3Coo-)
Titration of Amino Acid
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Zwitterion is isoelectric (pI) form with zero charge
pI is the average of pK1 +pK2 /2
Isoelectric point: pH at which an amino acid is
electrically neutral, that is the sum of positive charge
equal the sum of the negative charges.
Titration Curve of Alanine
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At low pH both (CooH &
NH2) are protenated.
As pH of the solution
raised (CooH)
dissociated by donating
a proton in the medium
forming hydroxylate
group (Coo-) so the
molecule is dipolar form
called Zwitter ion or
Isoelectric forms
Application of Henderson-Hasselbalch
Equation
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Henderson-Hasselbalch Equation can be used
to calculate how the pH of the physiological
solution responds to change in the
concentration of week acids and /or salt
form.
Application of HendersonHasselbalch Equation
Bicarbonate as a buffer:
pH=pKa+ log [HCO3-]
[H2CO3]
 ↑ bicarbonate ↑ pH
↑ CO2 as in pulmonary obstruction
 pH
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Drug Absorption
Drugs are either weak
acid or weak bases & the
uncharged form is more
permeable through
Membranes.
Acid: HA  H+ + ABase: BH+  B + H+
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Drug Absorption
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however, acid drugs permeate more in
low pH whereas basic drugs permeate
more in high pH
Example: Aspirin
Buffering occurs within ± pH unit of the
pka where [A-]=[HA]
Activity 1
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Classify each of the 20 amino acids
according to the side chain on the a
carbon as non-polar, polar, sulfurcontaining, basic, acidic, or amide
derivative
Primary Structure of Proteins
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How is protein formed?
By peptide bond
Character of peptide bond:
Amide bond between α-COOH & α-NH2
Not broken by heating or urea
Can be broken by strong acids or base
at high temp. or by proteolytic enzymes
Peptide bond
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AA are joined covalently by peptide bond
which are amide linkage between α-carboxyl
group of one amino acid and α-amino group
of another.
Partial double bond
Rigid & planar
Uncharged but polar
-NH & -C=O groups are involved in hydrogen
bonding
Amino acids can be linked by peptide bonds
Peptide bond
- condensation reaction (loss of one H2O)
-Several amino acids can be linked, forming a polypeptide chain
- backbone: -NH2, a-Carbon, -COOH ; side chains protrude from backbone
- Convention: amino terminus taken as beginning of polypeptide: N  C
Side chains
Peptide
backbone
Side chains
Peptide bonds may be cis or trans
configured
The planar arrangement of the atoms in the peptide bond (which is required for resonance)
can be realised in two ways:
Trans:
Carbonyl-O and amide-H on different sides of the peptide group
Cis:
Carbonyl-O and amide-H on the same side of the peptide group
Usually, the trans configuration is strongly favoured, since there is no steric hindrance
H
C–N
O
O
H
C–N
Peptides – polarity N  C
N-terminus
(amino end)
C-terminus
(carboxyl end)
Naming the peptide
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Free NH2 on left is N-terminal & free COOH on
the right is C-terminal
All proteins are read from N- to C- terminal
end of peptide
Polypeptide is linkage of many AA via peptide
bonds
Each AA in a polypeptide is called a residue or
moiety
Ex: val-gly-leu is called valylglycylleucine
Levels of protein Structure
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Primary structure: sequence of AA in a
polypeptide chain. Important in genetic
disease.
Secondary structure: regular arrangements of
AA in linear sequence (Ex.α-helix ,β-pleated
sheet & B-bend).It describe the geometrical
arrangment of polypeptide bond around one
axis.
Tertiary structure: folding of protein domains
of a single polypeptide chain in three
dimensional structure.
Quaternary: more than one polypeptide
chain.
Secondary Structure of Protein
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1. α-helix:
Spiral structure
Coiled polypeptide backbone core with
the side chains of AA extending
outward from central axis to avoid
sterric hindrous
Stability of α-helix is by hydrogen bond
formation between -C=O----HN-(
Carbonyl O2 and amide H2
Secondary Structure of Protein
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Each turn of α-helix contains 3:6 of AA
AAs that disrupt α-helix are:
Pro (insert a kink)
Charged AA e.g glu,asp,his,lys, arg
(form ionic bonds)
Bulk side chain AAs e.g trp,val,Ile
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Peptide backbone
N-C-C-N
Secondary Structure of Protein
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β-Sheet
All of peptide bonds are involved in H.B and,
therefore, is fully extended.
Appears pleated & as arrows and can be
formed from two or more separate
polypeptide chains.
H.B is perpendicular to polypeptide backbone
& called interchain bonds of separate
polypeptide chains or intrachain of a single
polypeptide chain
Types of β-Sheet
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Pararllel & antiparallel
Compare between
α-helix & β-sheet
Secondary Structure of Protein
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3. B-Bends
B-bends (reverse turns)
B-bends reverse the direction of a
polypeptide chain helping it form a compact
globular shape
Found on surface of protein molecule
Often include charged residues
Generally of four AAs one is pro that form
kink and gly
Stabilized by H.B & ionic bonds
4. Nonrepetitive secondary
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Half of globular protein is organized as
repetitive structure such as α-helix and
β-sheet, the remainder is formed as
loop or coil conformation called
nonrepetitive secondary structure
Nonrepetitive secondary structure are
not “random”but have less regular
structure than α-helix, β-sheet & Bbends
Supersecondary Strucure
(Motifs)
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Secodary structures, such as α-helix, βsheet, nonrepetitive & B-bend, are
elements to form the core region
(interior of protein mol) and at the
surface of protein mol loop regions (Bbends) are the connectors to the core
forming the motifs
Tertiary Structure
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Folding of basic units (domain) to form a final
arrangement of a single polypeptide chain
(monomeric protein)
Domain is a functional three dimensional
structure units of a polypeptide
Polypeptide chains > 200 AAs consist of 2 or
more domains
Core of a domain is built from different motifs
(supersecondary structure)
Interactions stabilizing Tertiary
Structure
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Interactions between side chains of AAs
determine the way a polypeptide fold to form
the compact structure
Types of interactions that form tertiary
structure:
1. Disulfide bond, covalent strong bond
2. Hydrophobic bond, nonpolar side chains
3. Hydrogen bond, between N or O & H
4. Ionic bond, between - & + groups
Interactions stabilizing Tertiary
Structure
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Protein folding: Interaction between the
side chains of AA determine how long
polypeptide chain folds into the intricate
three-dimensional shape of the
functional protein.
As a peptide folds, its AA side chains
are attracted and repulsed according to
their chemicals properties.
Protein Folding
Trial & error of side chain
interactions seek the
configurations of a protein
with a low energy state.
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Chaperones in Protein Folding
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Chaperones are specialized gp of
proteins required for proper folding of
proteins
Chaperones also called “heat shock”
proteins
Chaperones interact with polypeptide at
various stages during folding process
Functions of Chaperones
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Act as unfold unit until synthesis is
finished
Act as catalysts in final stages of folding
Protect proteins as they fold
Quaternary Structure of Protein
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Proteins that consist of more than one
polypeptide chain (multimeric)
If a protein consists of two subunits,
protein is dimeric. If three, protein is
trimeric
Subunits are connected by noncovalent
interactions (H.B, ionic & hydrophobic)
Subunits may work independently or
cooperatively (Hb)
Denaturation of Proteins
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Unfolding & disorganization of proteins'
secondary and tertiary structures
without hydrolysis of peptide bonds
Denaturing agents are: heat, organic
solvents, mechanical mixing, acids &
bases, detergents & heavy metals (Pb,
Hg)
Denatured proteins are insoluble &
precipitated from solution
Protein Misfolding
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Misfolding of proteins may occurs
spontaneously or be caused by a mutation in
a particular gene, which then produced an
alter protein.
In addition, some normal proteins can after
abnormal proteolytic cleavage, take on a
unique conformational state that leads to the
formation of long , fibrillar protein
assemblies consists of β-pleated sheets
Accumulation of these spontaneously
aggregating proteins called amyloids
Protein Misfolding
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If proteins are misfolded (improper folding) & not
degraded, these proteins may be deposited & cause
diseases called amyloidoses
Alzheimer disease is a degenerative disease caused
by accumulation of misfolded proteins (amyloid
plaque)
Alzheimer disease: normal proteins after abnormal
chemicals processing , take on unique conformational
state that leads to formation of neurotoxin amyloid
proteins assemblies consisting of β-pleated sheets.
Protein Misfolding
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Prion disease: It has been strongly
implicated as the causative agent of
transmissible spongiform encephalopathies ( TSEs)
Creutzfeldt-Jacob disease in humans
Mad caw disease in cattle.
In TSEs, the infective agent is an altered version of a
normal prion protein that act as template for
converting normal protein to the pathogenic
conformation.
Activity 2
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Define primary, secondary and tertiary
structure of proteins, mention the
bonds that stabilize each structure.