Proteins: Function & Structure
Proteins
1. Cellular Overview
1. Functions
2. Key Properties
2. Core Topics
1. Amino Acids: properties, classifications, pI
2. Primary Structure, Secondary Structure, and
Motifs
3. Tertiary Structure
1. Fibrous vs. Globular
4. Quaternary Structure
Amazing Proteins: Function
1. Catalysts (Enzymes)
•The largest class of proteins, accelerate rates of reactions
DNA Polymerase
CK2 Kinase
Catalase
2. Transport & Storage
Hemoglobin
Serum albumin
Ion channels
Ovalbumin
Casein
Amazing Proteins: Function
4. Structural
Collagen
Keratin
Silk Fibroin
5. Generate Movement
Actin
Myosin
Amazing Proteins: Function
5. Regulation of Metabolism and Gene Expression
Lac repressor
Insulin
6. Protection
Immunoglobulins
Thrombin and
Fibrinogen
Venom Proteins
Ricin
Amazing Proteins: Function
7. Signaling and response (inter and intracellular)
Apoptosis
Membrane proteins
Signal transduction
Amazing Proteins: Properties
• Biopolymers of amino
acids
• Contains a wide range of
functional groups
• Can interact with other
proteins or other
biological macromolecules
to form complex
assemblies
• Some are rigid while
others display limited
flexibility
a-Amino Acids: Protein Building Blocks
R-group or side-chain
R
a-amino group
+
H3N
C
C
-
H
a-carbon
O
O
Carboxyl group
Amino acids are zwitterionic
• “Zwitter” = “hybrid” in German
R1
H2N
C
H
R1
O
C
H3 N
OH
+
C
O
C
-
H
O
• Fully protonated forms will have specific pKa’s
for the different ionizable protons
• Amino acids are amphoteric (both acid and base)
Stereochemistry of amino acids
Stereochemistry of amino acids (AA)
• AA’s synthesized in the lab are racemic mixtures.
AA’s from nature are “L” isomers
• These are all optically-active except for glycine
(why?)
Synthesis of Proteins
R1
+
H3N
C
C
-
H
R2
O
+
+
H3N
O
C
R1 O
H3N
C
H
C
-
H
+
O
C
O
peptide
bond
R2
NH C
O
+ H2O
C
-
H
O
Synthesis of Proteins
R1 O
H 3N
+
C
C
R2
NH C
C
-
H
R3
O
+
H3N
O
H
H 3N
C
H
C
C
C
-
H
R1 O
+
+
O
O
R2 O
NH
C
H
R3
C
NH
C
O
C
-
H
O
Synthesis of Proteins
R1 O
H 3N
+
C
C
R2 O
NH
H
C
R3
C
NH
H
C
C
C
H
N-Terminal End
+
H3N
C
O
C
-
-
O
O
H
R2 O
R1 O
H 3N
+
C
H
+
R4
O
NH
C
H
C
NH
R3
O
C
C
H
R4
NH
C
O
C
-
H
O
C-Terminal End
Synthesis of Proteins
R2 O
R1 O
H3 N
+
C
C
NH
C
C
NH
O
C
C
R4
NH
C
O
C
-
H
H
H
R3
H
O
R1
O
=
≠
R4 O
+
H3N
C
C
R2 O
NH
C
C
NH
R3
O
C
C
NH
C
C
-
H
H
H
H
O
Synthesis of Proteins
R2 O
R1 O
H3 N
+
C
C
NH
C
C
NH
O
C
C
R4
NH
C
O
C
-
H
H
H
R3
H
O
=
≠
R3 O
R1 O
H3 N
+
C
H
C
NH
C
H
C
NH
R2
O
R4
C
C NH
C
H
O
C
-
H
O
Synthesis of Proteins
R2 O
R1 O
H3 N
+
C
C
NH
C
C
NH
O
C
C
R4
NH
+
H3N
C
C
R2 O
NH
C
C
NH
C
O
H
≠
R4 O
C
O
-
H
H
H
R3
R3
O
C
C
R1
NH
C
O
C
-
H
H
H
O
H
≠
R3 O
R1 O
H3 N
+
C
H
C
NH
C
H
C
NH
R2
O
R4
C
C NH
C
H
O
C
-
H
O
COMMON AMINO ACIDS
20 common amino acids make up the multitude
of proteins we know of
Amino Acids With Aliphatic Side Chains
Amino Acids With Aliphatic Side Chains
Amino Acids With Aliphatic Side Chains
Amino Acids With Aromatic Side Chains
Amino Acids with Aromatic Side Chains Can Be
Analyzed by UV Spectroscopy
Amino Acids With Hydroxyl Side Chains
Amino Acid with a Sulfhydryl Side Chain
Disulfide Bond Formation
Amino Acids With Basic Side Chains
Amino Acids With Acidic Side Chains and Their
Amide Derivatives
There are some important uncommon amino acids
pH and Amino Acids
Net charge: +1
Net charge: 0
Net charge: -1
Characteristics of Acidic and Basic Amino Acids
• Acidic amino acids
▫ Low pKa
▫ Negatively charged at
physiological pH
▫ Side chains with –COOH
▫ Predominantly in
unprotonated form
• Basic amino acids
▫ High pKa
▫ Function as bases at
physiological pH
▫ Side chains with N
Isoelectic point (pI)
• the pH at which the compound is electrically
neutral
▫ Equal number of (+) and (-) charge
• At pH < pI
• At pH > pI
amino acid is (+)
amino acid is (-)
• CRITICAL FOR: protein analysis, purification,
isolation, crystallization
We use different “levels” to fully describe the structure of a
protein.
Primary Structure
• Amino acid sequence
• Standard: Left to Right means N to C-terminal
• Eg. Insulin (AAA40590)
MAPWMHLLTVLALLALWGPNSVQAYSSQHLCG
SNLVEALYMTCGRSGFYRPHDRRELEDLQVEQ
AELGLEAGGLQPSALEMILQKRGIVDQCCNNI
CTFNQLQNYCNVP
• The info needed for further folding is contained
in the 1o structure.
Secondary Structure
• The regular local structure based on the
hydrogen bonding pattern of the polypeptide
backbone
▫ α helices
▫ β strands (β sheets)
▫ Turns and Loops
• WHY will there be localized folding and
twisting? Are all conformations possible?
Consequences of the Amide Plane
Two degrees of freedom per residue for the
peptide chain
• Angle about the C(alpha)-N bond is denoted phi
• Angle about the C(alpha)-C bond is denoted psi
• The entire path of the peptide backbone is
known if all phi and psi angles are specified
• Some values of phi and psi are more likely than
others.
The angles phi and psi are
shown here
See blackboard for
explanation why the
peptide bond is planar
Unfavorable orbital overlap precludes some
combinations of phi and psi
• phi = 0, psi = 180 is unfavorable
• phi = 180, psi = 0 is unfavorable
• phi = 0, psi = 0 is unfavorable
Steric Constraints on phi & psi
Sasisekharan
• G. N. Ramachandran was
the first to demonstrate
the convenience of
plotting phi,psi
combinations from known
protein structures
• The sterically favorable
combinations are the
basis for preferred
secondary structures
α Helix
•First proposed by Linus Pauling and Robert Corey in 1951.
•3.6 residues per turn, 1.5 Angstroms rise per residue
•Residues face outward
α Helix
• α-helix is stabilized by H-bonding
between CO and NH groups
• Except for amino acid residues at the end
of the α-helix, all main chain CO and NH
are H-bonded
α Helix representation
β strand
• Fully extended
• β sheets are formed by linking 2 or more strands
by H-bonding
• Beta-sheet also proposed by Corey and Pauling
in 1951.
PARALLEL
ANTIPARALLEL
The Beta Turn
(aka beta bend, tight turn)
•allows the peptide chain to reverse
direction
•carbonyl C of one residue is H-bonded
to the amide proton of a residue three
residues away
•proline and glycine are prevalent in beta
turns
Mixed β Sheets
Twisted β Sheets
 Loops
What Determines the Secondary Structure?
• The amino acid sequence determines the
secondary structure
• The α helix can be regarded as the default
conformation
– Amino acids that favor α helices:
Glu, Gln, Met, Ala, Leu
– Amino acids that disrupt α helices:
Val, Thr, Ile, Ser, Asx, Pro
What Determines the Secondary Structure?
• Branching at the β-carbon, such as in valine,
destabilizes the α helix because of steric
interactions
• Ser, Asp, and Asn tend to disrupt α helices
because their side chains compete for H-bonding
with the main chain amide NH and carbonyl
• Proline tends to disrupt both α helices and β
sheets
• Glycine readily fits in all structures thus it does
not favor α helices in particular
Can the Secondary Structure Be Predicted?
• Predictions of secondary structure of proteins
adopted by a sequence of six or fewer residues
have proved to be 60 to 70% accurate
• Many protein chemists have tried to predict
structure based on sequence
▫ Chou-Fasman: each amino acid is assigned a
"propensity" for forming helices or sheets
▫ Chou-Fasman is only modestly successful and doesn't
predict how sheets and helices arrange
▫ George Rose may be much closer to solving the
problem. See Proteins 22, 81-99 (1995)
Modeling protein folding with
Linus (George Rose)
Tertiary Structure
• The overall 3-D fold of the polypeptide chain
• The amino acid sequence determines the
tertiary structure (Christian Anfinsen)
• The polypeptide chain folds so that its
hydrophobic side chains are buried and its polar
charged chains are on the surface
▫ Exception : membrane proteins
▫ Reverse : hydrophobic out, hydrophilic in
• A single polypeptide chain may have several
folding domains
• Stabilized by H-bonding, LDF, noncovalent
interactions, dipole interactions, ionic
interactions, disulfide bonds
Fibrous and Globular Proteins
Fibrous Proteins
• Much or most of the polypeptide chain is
organized approximately parallel to a
single axis
• Fibrous proteins are often mechanically
strong
• Fibrous proteins are usually insoluble
• Usually play a structural role in nature
Examples of Fibrous Proteins
• Alpha Keratin: hair, nails, claws, horns, beaks
• Beta Keratin: silk fibers (alternating Gly-Ala-Ser)
Examples of Fibrous Proteins
• Collagen: connective tissuetendons, cartilage, bones,
teeth
▫ Nearly one residue out of
three is Gly
▫ Proline content is unusually
high
▫ Unusual amino acids found:
(4-hydroxyproline, 3hydroxyproline , 5hydroxylysine)
▫ Special uncommon triple
helix!
Globular Proteins
• Most polar residues face the outside of the
protein and interact with solvent
• Most hydrophobic residues face the interior of
the protein and interact with each other
• Packing of residues is close but empty spaces
exist in the form of small cavities
• Helices and sheets often pack in layers
• Hydrophobic residues are sandwiched between
the layers
• Outside layers are covered with mostly polar
residues that interact favorably with solvent
An amphiphilic helix in
flavodoxin:
A nonpolar helix in citrate
synthase:
A polar helix in calmodulin:
Quaternary Structures
• Spatial arrangement of subunits and the nature
of their interactions. Can be hetero and/or
homosubunits
• Simplest example: dimer (e.g. insulin)
ADVANTAGES of 4o Structures
▫
▫
▫
▫
Stability: reduction of surface to volume ratio
Genetic economy and efficiency
Bringing catalytic sites together
Cooperativity
Protein Folding
• The largest favorable
contribution to folding
is the entropy term for
the interaction of
nonpolar residues with
the solvent
• CHAPERONES
assist protein
folding
▫ to protect nascent
proteins from the
concentrated protein
matrix in the cell and
perhaps to accelerate
slow steps