Download 2012 jf lecture 2.pptx

Proteins Dr. Sarah Doyle School of Biochemistry and Immunology Lecture 2
Protein conformation
The Genetic Code
The Genetic Code:
Genes are made up of sets of 3
nucleotides (codons) which
encode a sequence of amino
acids that form a polypeptide
DNA codes for RNA
(according to the base pair rule)
RNA codes for protein:
3 nucleotides encode 1 amino acid:
Protein:
A sequence of amino acids form a
polypeptide
The genetic code
???How many different proteins are there in a human cell??? -­‐ Latest es@mate: ~25,000 different proteins occur in humans -­‐ How many proteins are needed to make a single cell? -­‐ Different for different species: -­‐ Worm (nematode): 19,000 -­‐ Fly (fruit fly): 16,000 -­‐ Yeast: 5,000 -­‐ Mycobacteria: 500 -­‐ Hepa@@s C virus: 8 (uses cell it infects) What is the structure of a protein?
• Each of the ~25,000 human proteins have
specific structure and function
• Protein shape is very important to its function
proteins interact with other proteins
Eg. antibody
• Diverse functions = diverse structures
• Unique function = unique 3D shape
• Protein Shape=Conformation
Protein stuctures
Polypeptides
•  Proteins
–  Are polymers of amino acids (aa)
aa-aa-aa-aa-aa-aa-aa-aa-aa-aa-aa-aa-aa-aa
polymer of aa = polypeptide
•  A protein
–  Consists of one or more polypeptides
polypeptide(s) = protein
• 1949: Fred Sanger determined the sequence of insulin
Amino Acid Monomers
•  Amino acids
–  Are organic molecules possessing both carboxyl and
amino groups
–  Differ in their properties due to differing side chains,
called R groups
R= reactive group
Backbone of each aa –  20 different R groups = 20 different amino acids
Amino Acid Monomers Unionised form
Zwitterion form
• internal transfer of a H ion from the -COOH to the -NH2 group
= an ion with both a negative charge and a positive charge
= over electrically neutral
4 subgroups of Amino acid
-­‐ There are 20 naturally-­‐occurring amino acids -­‐ Grouped into 4 subgroups, depending on the R-­‐group: -­‐ Non-­‐polar: hydrophobic -­‐Polar + Neutral: hydrophilic -­‐ Polar + Acidic: hydrophilic hydrophilic
hydrophobic
-­‐ Polar + Basic: hydrophilic A Polar compound is one where the spread of charge is not even, with some
atoms carrying a partial negative charge and others a partial positive charge
1. Non-polar amino acids
hydrophobic / water insoluble
2. Polar amino acids
hydrophilic / water soluble
Electrically charged amino acids
water soluble
3. Acidic
4.Basic
Amino acid abbreviations
Non-polar
Polar
AMINO ACID glycine THREE LETTER CODE Gly SINGLE LETTER CODE G alanine Ala A valine Val V leucine Leu L isoleucine Ile I methionine phenylalanine Met Phe M F tryptophan Trp W proline Pro P AMINO ACID serine threonine cysteine tyrosine asparagine glutamine THREE LETTER CODE Ser Thr Cys Tyr Asn Gln SINGLE LETTER CODE S T C Y N Q Neutral
AMINO ACID aspar@c acid glutamic acid THREE LETTER CODE Asp Glu SINGLE LETTER CODE D E Acidic
AMINO ACID lysine arginine his@dine THREE LETTER CODE Lys Arg His SINGLE LETTER CODE K R H Neutral
Basic
Essential amino acids
-those amino acids that an organism can not synthesize on its own
Essen@al Nonessen@al Isoleucine Alanine Leucine Arginine* Lysine Aspartate Methionine Cysteine* Phenylalanine Glutamate Threonine Glutamine* Tryptophan Glycine* Valine Proline* His@dine Serine* Tyrosine* Asparagine* Obtained from diet
* Conditionally essential, for some individuals, eg tyrosine in PKU
Phenylketonuria (PKU)
•  Classical PKU is caused by a mutated gene for the
enzyme phenylalanine hydroxylase (PAH)
•  PAH converts the essential amino acid phenylalanine to
other essential amino acids eg tyrosine.
•  Lack of PAH causes phenylalanine accumulation, lack of
tyrosine – normal at birth with gradual loss of mental
function, severe learning disabilities and seizures
•  Treatment: intake of phenylalanine low, supplement diet
with tyrosine
Heel prick
“Guthrie test”
Amino Acid Polymers
•  Amino acids - Are linked by peptide bonds
Peptide
bond
OH
OH
SH
CH2
CH2
H
H
N
Peptide bond is formed by a
dehydration reactionCatalytic reaction
CH2
H
C
C
H
O
N
H
C
C
H
O
OH H
(a)
N
C
C
H
O
OH
H 2O
OH
CH2
Peptide bond - covalent bond
H
H
H N
(b) C
C
H
O
Amino end
(N-terminus)
Side chains
SH
Peptide
CH2 bond CH2
OH
N
H
C
C
H
O
N
C
C
H
O
Carboxyl end (C-­‐terminus) OH
Backbone
Protein Conformation and Function
•  Polypeptides
- formed one at a time starting from N-terminus
- range from a few monomers to 1000 or more
•  Specific polypeptides- unique sequence of aa’s (as determined by the genetic code)
•  Sequence of the aa polymer determines the 3D shape of the
polypeptide
•  Proteins are not just chains of aa’s, they are defined by their
shape – interactions between backbone residues and R-groups
•  A protein’s specific conformation - determines how it functions
eg. Enzyme binds substrate
The function of a protein is
inextricably linked to its shape
hemoglobin
IgG
Adenylate kinase
insulin
Glutamine synthetase
Molecular surface of several proteins showing their compari@ve sizes •  Two models of protein conformation
Groove
Eg. Lysozyme- an enzyme that
Breaks down bacterial cell walls
by recognizing and binding to
specific molecules on the bacteria
(a) A ribbon model
Groove
(b) A space-filling model
Four Levels of Protein Structure
•  Primary structure
–  Is the unique sequence of amino acids in
a polypeptide
HN
Amino acid
+
Gly Pro Thr Gly
Thr
Gly
3
Amino
end
subunits
Glu
CysLysSeu
LeuPro
Met
Val
Lys
Val
Leu
Asp
AlaVal Arg Gly
Ser
Pro
Ala
aa sequence determined by
inherited genetic information
Glu Lle
Asp
Thr
Lys
Ser
Lys Trp Tyr
Leu Ala
Gly
lle
Ser
ProPheHis
Glu
Ala Thr PheVal
Asn
His
Ala
Glu
Val
Thr
Asp
Tyr
Arg
Ser
Arg
Gly Pro
Thr Ser
Tyr
lle
Ala
Ala
Leu
Leu
Ser
Pro
SerTyr
Thr
Ala
Val
Val
Glu
ThrAsnProLys
o
c –
o
Carboxyl end
Eg lysozyme is 129aa long
•  Secondary structure
–  Is the folding or coiling of the polypeptide into a
repeating configuration
–  Includes the α helix and the β pleated sheet
–  Result of Hydrogen bonding between the
repeating backbone of a polypeptide
β pleated sheet
O H H
C C N
Amino acid
subunits
C N
H
R
R
O H H
C C N
C C N
O H H
R
C C N
OH H
R
O
R
C
H
R
O C
O C
N H
N H
N H
O C
O C
H C R H C R
H C R H C
R
N H O C
N H
O C
O C
H
C
O
N H
N
C
C
R
R
C
C
H
R
O H H
C C N
C C N
OH H
R
O
C
H
H
H C N HC
C
C N HC N
N
H
H
C
O
C
C
O
R
R
O
N
O H H
C C N
R
R
H
R
O
C
H
H
NH C N
C
H
O C
R
R
C C
O
R
H
C
N HC N
H
O C
H
α helix
-Repeated coils or folds in patterns contribute to the
overall conformation of a protein
Hydrogen bonds
•  Hydrogen bonds are formed by the attraction between a partial
positive charge on the H atom of the amino group and the partial
negative charge on the O atom of the peptide bond
•  Alpha helix - the bonds are formed
between repeating atoms on the
same polypeptide chain
eg collagen
•  Beta sheets – the bonds are formed
between polypeptide chains
lying side by side
eg. Silk fibrion
Individually weak, but strong when repeated
R
aa
aa
Δ+
C
C
ΔN
Δ-O
H
HΔ+
Δ+
ΔH O
H
N
Δ-
C
R
C
Δ+
aa
aa
Alpha-helix
Beta pleated sheet
Secondary structure
Beta sheet
Alpha helix
Results from interactions between atoms in the polypeptide backbone
Tertiary structure
–  Is the overall three-dimensional shape of a
polypeptide ; final shape of a polypeptide
–  Results from interactions between the side
chains of the amino acids
Hydrogen
bond
CH2 CH
2
O
H
O
H 3C
CH
CH3
H 3C
CH3
CH
Hydrophobic
interactions and
van der Waals
interactions
HO C
CH2
CH2 S S CH2
Disulfide bridge O
CH2 NH3+ -O C CH2
Ionic bond
Polypeptide
backbone
Tertiary structure
•  Hydrophobic interactions:
Non-polar “R” groups are repelled by water
They tend to cluster together
Force themselves into the core of a protein
Stabilize the overall structure
Tertiary structure
•  Van der Waals interactions:
occur between hydrophobic non-polar
side chains in close contact
Valine
H 3C
CH
CH3
H 3C
CH3
CH
Hydrophobic
interactions and
van der Waals
interactions
Polypeptide
backbone
Tertiary Structure
•  Hydrogen bonds – between polar side
chains
•  Examples of amino acid side chains
that may hydrogen bond to each other:
Two alcohols: ser, thr, and tyr.
Alcohol (OH) and an acid : asp and tyr
Two acids (COO-): asp and glu
Alcohol and amine (NH3+): ser and lys
Alcohol and amide (NH2): ser and asn
Tertiary structure
•  Ionic bonds /Salt bridges
form between positively and
negatively charged side groups
•  result from the neutralization of
an acid and amine on side
chains.
•  The final interaction is ionic
between the positive
ammonium group and the
negative acid group.
•  Any combination of the various
acidic or amine amino acid
side chains will have this
effect.
Tertiary structure
•  Disulphide bridges form
between 2 cysteine residues
•  Disulfide bonds are formed by
oxidation of the sulfhydryl
groups (-SH) on cysteine.
•  Different protein chains or
loops within a single chain are
held together by the strong
covalent disulfide bonds.
•  Eg. Insulin contains important
disulphide bridges.
Disulfide bonds
keratin
Quaternary Structure
–  Is the overall protein structure that results from the
aggregation of two or more polypeptide subunits
–  A variety of bonding interactions including hydrogen
bonding, salt bridges, and disulfide bonds hold the
various chains into a particular geometry.
–  There are two major categories of proteins with
quaternary structure - fibrous and globular.
Quaternary structure
Fibrous
Trimer of alpha-helical
Polypeptide chains
Globular
Globular protein4 Polypeptide chains/ subunits
Primarily alpha-helical
β Chains
Iron
Heme
α Chains
Collagen
Structure allows for great strength
Rigid resistant to stretch
Function = connective tissue in
skin, bone, tendons, ligaments.
40% of all human protein
Hemoglobin
4 polypeptides bind together to
form a round globular shape
Functions = carries oxygen
Collagen
Hemoglobin
Quaternary structure Fibrous
Globular
ribonuclease
Beta sheet structure of silk fibroin
allows for strength and flexibility
Globular proteins are usually
round in shape and tend to be
a mix of alpha-helical and Beta
sheet
Determining Protein Structure
•  X-­‐ray crystallography –  Is used to determine a protein’s three-­‐
X-ray
dimensional structure diffraction
pattern
Photographic film
Diffracted X-rays
X-ray
X-ray
beam
source
Crystal Nucleic acid Protein
(a) X-ray diffraction pattern
(b) 3D computer model
Protein structure summary…
•  The conformation of a protein determines its function
•  Proteins are made of polypeptides which are polymers
of amino acids
•  Amino acid polymers are linked by peptide bonds
•  The amino acid sequence determines the 3-D shape
of the Protein
•  There are four levels of protein structure
Primary Secondary
Ter@ary Quaternary