Section Week 3 Junaid Malek, M.D.
Section Week 3Junaid Malek, M.D.
Biological Polymers• DNA
• 4 monomers (building blocks), limited structure (double-helix)
• RNA
• 4 monomers, greater flexibility, multiple structures
• Proteins
• 20 Amino Acids, greatest variety of possible structures
Proteins
• 4 major functions
• Structural
• Enzymatic
• Carriers
• Regulatory
Amino Acids: Components
• Major components include amine group, carboxylate group and the side chain
• Remember that 19 of 20 amino acids are chiral
• Amine and carboxylate groups participate in peptide linkages
Amino Acids: Structure
• Important to memorize structures, 3-letter and 1-letter codes for the exam
The Peptide Bond
H3N O
OH3N O
O
OH
+
H3N NH
O
O
O
OH
H2O
alanine serine
peptide (amide) bond in alanylserine
H3N NH
O
O
O
OHOH2
Double bond property of a peptide bond
H2NNH
O
O
OH
OH
H2NHN
O
O
OH
OH
• The reason why a peptide bond is not capable of rotating
• Rotation also restricted by R groups
cis vs. trans nomenclature
H2NN
O
R
R'
O
OH
H2NN
O
R R' O
OHH
H
Trans: The α-carbonsare on opposite sides
H2N
N
O
R
R'
O
OH
H2N
N
O
RR'
O
OH
H
H
Cis: The α-carbonsare on the same side
Why is trans favored over cis?
• Trans configuration favored due to less steric hindrance between R side-chain and peptide backbone
N
O
ON
O
OH
H
Heavily favored
Trans Cis
Glycine• Only achiral amino acid
• Adds flexibility to any polypeptide chain
• Less steric hindrance
Proline
• The only exception to the cis vs. trans rule
• Because the nitrogen contains two substituents, both cis and trans contain steric strain. Therefore, they exist in nearly equal proportions.
Slightly favored
Trans Cis
NC
OON
O
CO
Cysteine
• Sulfur-containing amino acid
• Capable of undergoing reduction-oxidation reactions
• Don’t need to know specifics of red-ox reactions
• Just know that two distinct states exist that are not spontaneously interconvertable
• Disulfide bonds are important for protein folding
Histidine• Can act as an acid or a
base
• Note that pKa is near physiological pH
Draw the following oligopeptide at pH=7
• Lys-Phe-Met-Arg
H3N+
O
NH
+NH3
HN
O
S
NH
O
O-
O
NH
NH2
H2N+
Lys
Phe
Met
Arg
H3N+
O
NH
H2N
HN
O
NH
O
O-
O
OH
Gln
Ile
Glu
Thr
O
-O
O
Draw the following oligopeptide at pH=7
• Gln-Ile-Glu-Thr
H3N+
O
NH
+NH3
HN
O
S
NH
O
O-
O
NH
NH2
H2N+
Lys
Phe
Met
Arg
H3N+
O
NH
H2N
HN
O
NH
O
O-
O
OH
Gln
Ile
Glu
Thr
O
-O
O
Circle the Hydrophobic GroupsPut a square around the alcohol group
H3N+
O
NH
+NH3
HN
O
S
NH
O
O-
O
NH
NH2
H2N+
Lys
Phe
Met
Arg
H3N+
O
NH
H2N
HN
O
NH
O
O-
O
OH
Gln
Ile
Glu
Thr
O
-O
O
More Questions
H3N+
O
NH
+NH3
HN
O
S
NH
O
O-
O
NH
NH2
H2N+
Lys
Phe
Met
Arg
H3N+
O
NH
H2N
HN
O
NH
O
O-
O
OH
Gln
Ile
Glu
Thr
O
-O
O
• Are these oligopeptides chiral?
• Yes
• Which of the two peptides interacts best with DNA? Why?
• The peptide on the left, because it has two positively charged groups that can interact with negatively charged DNA
More Questions
• How many different pentapeptides can one make using the amino acids Gly, His, Cys, Ile, Try?
• 5!=5x4x3x2x1=120
• Which peptide would you expect to be more soluble in water, one rich in aspartate and lysine or one rich in valine and alanine?
• Aspartate and lysine have side-chains capable of H-bonding while valine and alanine do not
Protein Structure
• Primary Structure
• Secondary Structure
• Tertiary Structure
• Quaternary Structure
Primary Structure
• Linear sequence of amino acids bonded by peptide bonds
• No folding or side-chain interaction
• By convention, written from N-terminus to C-terminus
• As with DNA and RNA, directionality matters!
• The primary sequence of amino acids will determine how the protein folds
Secondary Structure• Local structures adopted by contiguous amino acids
• α-helix
• Helical corkscrew structure that is typically right-handed due to chirality of amino acids
• Carbonyl groups all point in same direction
• β-pleated sheet
• Form sheet-like structures represented by arrows. Origin of arrow indicates amino terminus while arrowhead indicates carboxyl terminus.
• Carbonyl groups point in alternating directions
α-helix: Myoglobin
β-sheet: Superoxide Dismutase
α-helix and β-sheet: Acetylcholinesterase
Tertiary Structure
• Folding of a single polypeptide chain
• Forms due to interactions among R-groups and secondary structures
• Allows for amino acids far away on the primary chain to lie in close proximity to one another
• All information allowing a protein to form tertiary structure can be found in the primary sequence
Quaternary structure
• Interaction between multiple tertiary structures (from distinct polypeptide chains)
• Classic example is hemoglobin
Why does a protein fold?
• Anfinsen experiment: does a primary structure determine protein folding?
• A protein (Ribonuclease A) was denatured using urea and reduced to break disulfide bonds between cysteine residues
• Observation made that when urea was removed, the denatured polypeptide chain folded back into a compact structure
• When protein oxidized, regained 90% of original function
Protein folding
• In a second experiment, the denatured polypeptide chain was first oxidized before removing urea
• Result was a non-functional protein trapped in a non-native conformation by incorrect pairing of disulfide bonds
• These bonds prevented the protein from achieving its native structure
The Anfinsen Experiment: Conclusion
• Given that we prevent premature oxidation (and hence incorrect disulfide bridging), the protein will “find” its lowest energy conformation
• Thus, all information for proper protein folding can be found in its primary sequence!
• This is in turn determined by RNA sequence, which is determined by DNA sequence
• We can therefore conclude that all complex protein folding can be determined by the sequence of just 4 different base pairs!!!
What drives protein folding?
• Thermodynamics!
• Hydrogen bonding along the backbone
• Hydrogen bonds of the R groups with each other or with the backbone
• Ionic interactions between the R groups
• Van der Waal’s interactions between R groups
• Disulfide bridges between cysteine residues
The Hydrophobic Effect• Tendency for non-polar substance to interact with
each other rather than with water
• This leads to burial of non-polar sidechains within the interior of a protein as it “collapses” to form a globular structure
• Driven by the energetically unfavorable situation of having water molecules organized and surrounding a non-polar molecule
• Driven also to a lesser extent by Van der Waal’s forces
Question: Protein Folding
• Segments of proteins that connect successive regions of secondary structures are referred to as “turns” or “bends”. These are often rich in glycine and proline residues. Why?
• Glycine is found in bends because of its small size
• Proline constrains the conformation a polypeptide chain can adopt, thus it is often the initiator of bends
Question: H-bonding and Secondary structure
• Indicate whether the following amino acids can form H-bonds to participate in the formation of α-helices or β-sheets
• Arginine
• Glutamate
• Glycine
• Phenlyalanine
• Proline
• Threonine
Question: Amino acid interactions
• Indicate the strongest interaction that can form between the side chains of the following amino acid pairs:
• Pro-Leu
• Van der Waal’s
• Arg-Thr
• Hydrogen bonding
• Ile-Val
• Van der Waal’s
Question: Amino acid interactions
• More amino acid pairs:
• Tyr-Phe
• Van der Waal’s
• Arg-Asp
• Ionic
• Cys-Met
• Van der Waal’s
Question: Amino acid interactions
• More amino acid pairs:
• Gly-Ala
• Van der Waal’s
• His-Glu
• Ionic
• Cys-Cys
• Disulfide bond
Gibbs Free Energy
Gibbs Free Energy• If ΔG is negative, the process is favored (energy is
released)
• Two components of free energy are enthalpy (H) and entropy (S)
• If ΔH is negative, the reaction (or system) is exothermic
• If ΔH is positive, the reaction is endothermic
• Remember your Second Law of Thermodynamics (the entropy of the universe is always increasing)
What can we say about ΔH and ΔS?
ΔH ΔS Reaction
(-) (+) Spontaneous
(+) (-) Non-Spontaneous
(-) (-) Temperature dependent
(+) (+) Temperature dependent
Gibbs Free Energy
• We can relate Keq and free energy using the equation ΔG°=-RTlnKeq
• R=Equilibrium constant
• T=Temp (Kelvin)
• At equilibrium, there is a relationship between the chemical equilibrium and the change in free energy (ΔG) that occurs as a result of the chemical reaction. The change in free energy is related to the natural log of the equilibrium constant.
• If ΔG is negative, the process is favored (energy is released)