Amino acids

1. How do you determine D/L config of amino acids?

a. In fisher projection, D/L is used based on the location of the amino group.
b. NH2 on the left (L isomer). NH2 on the right (D isomer). D/L are enantiomers
2. How many of the proteinogenic amino acids are chiral?
a. 20 proteinogenic but 19 are chiral: Glycine is Achiral
b. Absolute configuration (R/S), all are L and S except Cysteine which is R configuration
3. Is there a relationship between D/L or R/S configuration and +/-?
a. No clear relationship between D/L and rotation of plane-polarized light (+/-).
4. Provide the structure three-letter abbreviation, 1-letter code for the 20 common amino
acids, it is essential that you know that.
5. Define essential amino acids and let me know which amino acids are essential?
a. We cannot synthesize de novo but obtained from our diet
6. Which amino acids are hydrophobic or hydrophilic?
a. Hydrophobic: Nonpolar, alkyl/aliphatic, aromatic
b. Hydrophilic: Polar, neutral/-/+ charges
7. Which aa are hydrophobic and what are their characteristics
a. Glycine, Alanine, Valine, Isoleucine, Leucine (alkyl side chain), methionine and proline
b. Aromatic: Phenylalanine, tryptophan, and tyrosine (hydrophilic neutral, b/c or OH)
c. Aromatic R-groups can absorb UV light. The higher the concentration of protein in a
substance, the more UV light is absorbed
8. Which aa are hydrophilic and what are their characteristics?
a. Neutral: Serine, threonine, cysteine, asparagine, glutamine and tyrosine
b. Positive: Histidine (Found in active site, can lose or gain, close to ph= 7.0), Lysine and
Arginine
c. Negative: Aspartic and Glutamic (larger)
d. Found on surface of proteins and subject to them. Modifications (Phosphorylation), can
be oxidized or reduced, affecting conformation of protein
9. Define zwitterion/amphoteric.
a. Molecule with positive and negative charge with the net of zero
b. When a neutral amino acid is at physiological pH (pH = 7.4) → the pH > pKa-COOH,
therefore the carboxylic acid group becomes deprotonated. However, the pH < pKa-
NH2 → so the amino group remains protonated
10. What is pI and how do you determine it?
a. Isoelectric point is the pH at which an a.a’s zwitterion is the dominant species.
b. To determine pI, you need to calculate the midpoint between the +1->0 pka and the 0->-1
pka
11. What happens when a neutral aa is placed in an acidic or basic solution?
a. When a neutral amino acid is in acidic conditions (i.e. pH = 1) → both the amino and
carboxylic acid group will become protonated (since pH < pka) → therefore the overall
charge will be +1.
b. When a neutral AA is in basic conditions (pH=10 or above) → pH > pKa of both
functional groups → therefore both groups will be deprotonated → so the overall charge
of the amino acid is -1.
12. What does it mean by “basic” and “acidic” amino acids and how does this relate to pI?
a. A basic amino acid has an R group that is basic at neutral pH. Examples are histidine,
arginine, and lysine, their R groups tend to bind hydrogen protons so these residues are
often positively charged at neutral pH.
b. Acidic amino acids have R groups that tend to donate protons. Examples are glutamate
and aspartate, their R groups tend to donate protons so they are often negatively charged
 at neutral pH. The pI of the basic amino acids R groups is greater than 7 because they are
positively charged at neutral pH. Vice versa for the acidic amino acids, their pI is lower
than neutral pH so at neutral pH the negative deprotonated state predominates.
13. What are the pKas of the C and N-terminus, what do you think the pH of the R group in the
basic amino acids, acidic amino acids, cysteine, and tyrosine would be?
a. - Common side chain pKas: C terminus pKa = 2, N terminus = 9, cysteine pKr = 8.3,
tyrosine pKr = 10, histidine pKr = 6, lysine pKr = 10.5, Arginine pKr = 12.5, aspartate =
3.8, and glutamate = 4.3.
b. Since most amino acids with a neutral R group exist as a zwitterion at neutral pH you
should be able to reason that the COOH pka is far below 7 making it deprotonated and
that the NH2 pka is above 7 making it protonated. Since the basic amino acids are
positively charged at neutral pH that means their R group has a pka above 7, acidic amino
acids donate protons so their R group has a pka below 7 making it exist in the
deprotonated state. Cysteine and tyrosine are both protonated at neutral pH so their R
groups have pkas above 7.
14. Draw a peptide bond, know mechanism of peptide bond formation via condensation rxn
and hydrolysis
a. - Peptide bond formation = typical condensation and dehydration rxn which forms amide
bond
b. Amide bond is stabilized by resonance delocalization between the nitrogen lone pair and
the carbonyl oxygen → this gives the peptide bond partial double-bond character →
therefore rotation about this peptide bond is limited due to the peptide bond being planar
and rigid. 
15. How are peptide bond hydrolyzed?
a. Hydrolysis: acid catalyzed reaction in the presence of heat, non-specific (H30+)
b. Proteolytic enzyme: Specific e.g. trypsin (lysine and arginine)
16. Reaction for cysteine and its function
a. Cysteine is found in reducing conditions such as the intracellular environment of the
cell (Antioxidant). 
 However, when two cysteines are found in close proximity and are in oxidizing
conditions → the thiols become oxidized and a disulfide bridge is formed
between the two AAs → termed cystine. 
 Oxidizing conditions are found in the extracellular environment where
2 electrons and 2 protons are lost. 
b. This is important when certain proteins form their tertiary and quaternary structures as
it helps stabilize these protein
17. know the reactants involved in the synthesis of a-amino acids via the Strecker synthesis,
Gabriel malonic ester synthesis, and Hell-Volhard-Zelinsky reaction, where the R group,
carboxyl group, and amino groups come from
a. In Strecker Synthesis, an aldehyde/ketone (has the R group) reacts with an ammonium
chloride (donates the amino group) and cyanide salt to form the a-amino acid,
b. in the Gabriel malonic ester synthesis, a halogenated malonic ester (donates the carboxyl
group) reacts with phthalimide (donates the amino group) and an alkyl halide (donates
the R group) to form the a-amino acid,
c. in the Hell-Volhard-Zelinsky method you first generate an a-halocarboxylic (donates the
carboxyl group and R group) and then use ammonium (donates the amino group) in a
substitution reaction to form the a-amino acid.

Proteins structures
o Define primary, secondary, tertiary, and quaternary protein structure.
o Primary structure: This is the linear sequence of amino acids in a protein which determines
the secondary, tertiary and quaternary (if there is one) structure. This structure is
determined by covalent bonds (aka the peptide bonds) between amino acids. It is also the
location of the disulfide bond
o Secondary Structure: The secondary structure is the formation of folded structures → alpha
helices and beta-pleated sheets. Both secondary structures are dependent on backbone
intramolecular hydrogen bond interactions between the carbonyl oxygen and the amino
hydrogen. 
o The tertiary structure of proteins is the 3D folded conformation of the protein.  It is
determined by acid-base interactions, H-bonding, disulfide bridges, van der waal
interactions and hydrophobic packing. 
 Role of proline: check above in the amino acid section → already explained. 
 Role of cystine: formation of disulfide bridges between cysteine residues help
stabilize the polypeptide. 
 Hydrophobic bonding → hydrophobic residues in the presence of an aqueous
environment will prefer to be packing inside and not exposed  → whereas
hydrophilic residues will prefer to face the aqueous environment where they can
interact with the polar environment.
o Quaternary proteins: Protein with more than 1 polypeptide chain have a quaternary
structure.
 Example: hemoglobin → it is composed of 4 subunits each with a prosthetic group
called heme. 
 Conjugated proteins - proteins with prosthetic groups.
o The quaternary structure has several advantages brings catalytic sites closer together, can
induce cooperativity and allosteric effects, and reduces the amount of DNA needed to
encode the protein.  
o What is an a-helix, where does hydrogen bonding occur?
o Alpha helices are coiled structures that wrap around a central axis (Right Handed) → the
side chain of the amino acids point out of the alpha helix → example: keratin. 
 These are often found spanning the plasma membrane → e.g. in GPCRs (G-protein
coupled receptors). 
o are interactions between amino acids 3-4 residues apart attractive or repulsive (ex:
hydrophobicity/charge), propensity of a-helix to form in presence of adjacent bulky residues,
similarly charged residues, glycine, or proline, frequency of amino acids at the ends of the a-
helix due to the intrinsic dipole.
o Since amino acids located 3-4 residues apart from each other interact they are likely attracted
to each other through similar hydrophobicity and their hydrophobic interactions or opposite
charges and ionic interactions.
o Bulky residues and similarly charged residues decrease propensity of a-helix to form due to
steric hindrance or electrostatic repulsions. Glycine is too flexible and proline is too rigid
preventing formation of the a-helix. Intrinsic dipole of a-helix consists of a positive charge at
the n-terminus and negative charge at the c-terminus so oppositely charged amino acids exist
at those ends, ex: acidic amino acids which are negative at neutral pH would be at n-
terminus. 
o What is a B-sheet, where is hydrogen bonding, what is an antiparallel vs. parallel B-sheet, what
is a B-turn, are glycine or proline common to them?
o Beta-pleated sheets are rows of amino acids which lie parallel or antiparallel to one another
→ the side chains point above or below the plane → example: fibroin
o consists of adjacent peptide sequences with hydrogen bonds formed between the amino acids
that are across from one another. Parallel B-sheets have their n or c-termini pointed in the
same direction, antiparallel means they are in opposite directions, so adjacent peptides would
have n-terminus or c-terminus on one end. A b-turn is another secondary structure element
that consists of four amino acids that often connect the ends of beta-sheets. Glycine and
proline are common this element, the former due to its flexibility, the latter because when its
nitrogen is involved in peptide bonds it forms the cis isomer which is perfect for the 180
degree turn characteristic of the b-sheet.
o What is a super secondary structure aka motif, compare the structure, main roles, and water
solubility of fibrous vs. globular?
o A super secondary structure/motif describes the relationship between multiple secondary
structures such as beta-barrels which are between multiple beta-sheets, helix-turn-helix which
are between b-turns and alpha helixes, b-a-b loop which is between two beta-sheets and an
alpha helix, as well as zinc finger motifs which can bind DNA, RNA, or protein depending
on their subtype.
o Generalizations of globular vs. fibrous proteins. Globular proteins are spherical, functional
(ex: enzymes), water-soluble. fibrous proteins form long narrow strands.
o what is a protein domain, describe the theories of protein folding?
o A protein domain is a part of a protein that can evolve, function, and exist independently of
the rest of the protein
o If proteins randomly folded until the lowest energy native conformation was achieved this
would take forever since amino acids can technically exist in an infinite number of
conformations, so, there must be some sort of structure behind protein folding, and two
common theories provide explanations.
o What is the Levinthal paradox,
o If proteins randomly folded until the lowest energy native conformation was achieved this
would take forever since amino acids can technically exist in an infinite number of
conformations, so, there must be some sort of structure behind protein folding, and two
common theories provide explanations.
o describe the stepwise and molten globule theories of protein folding. What is the largest amino
acid and what is the average mass of an amino acid.
o The stepwise theory describes how nearby amino acids rearrange themselves until lower
energy secondary structures are achieved and then many different secondary structures can
rearrange to form lower energy motifs that then rearrange themselves to form lower energy
tertiary structures that can then rearrange themselves until the lowest energy quaternary
structure is formed.
o The second theory, the molten globule theory, says that the protein spontaneously collapses
to a lower energy molten globule that is characterized by hydrophobic interactions between
the interior residues and as the peripheral residues rearrange themselves the molten
globule traverses through many semistable intermediate states until it achieves its native
conformation. The stepwise and molten globule model ensure protein folding takes place in
 a reasonable time frame since they minimize the number of possible conformations amino
acids/proteins can exist in as the protein structure develops higher order structure
o Tryptophan is the largest amino acid. Average amino acid has mw of 110 da.

o Why is protein denaturation dangerous, how can temperature, pH, salt concentration, and
solvent can disrupt protein folding.
o Denaturation is the breakdown of the normal 3D conformation of the protein which can be affected by
temperature, pH, enzymes, detergents and chemicals. 
 Temperature: an increase in temperature → increases the kinetic energy in the polypeptide → this
kinetic energy ultimately becomes high enough to overcome the interactions holding the protein
together → denaturing the protein → therefore destroying the secondary, tertiary and quaternary
structure. 
 pH → breaks ionic bonds → therefore destroys the tertiary and quaternary structure.
 Chemicals → disrupt the hydrogen bonds within a protein → therefore destroying the secondary,
tertiary and quaternary structure.
 Enzymes → break the peptide bonds → therefore destroying the primary structure. 
 Detergents → solubilize the protein, breaking the non-covalent bonds → therefore destroying the
secondary, tertiary and quaternary structure.

Folding is the process by which a protein gains its proper 3D conformational shape and becomes active → more information
about this in the hydrophobic interactions and solvation section.

o When a hydrophobe is dropped in aqueous medium describe the enthalpy and entropy change,
o hydrophobic residues were on the surface of protein, then the water molecules would be forced to rearrange
themselves such that they maximize hydrogen bonding → but in doing so, they reduce entropy.
o the hydrophobic residues end up packing to the interior of the protein, to allow the hydrophilic residues to end up
on the surface which can interact with the solvation layer and therefore increase entropy → making solvation
energetically favourable → this process is called the hydrophobic effect. 
 a protein maximizes stability and This plays an important role in folding.
o What would a graph of %protein unfolding vs. increased temperature/salt concentration look
like and suggest about cooperativity?
o as temp/salt increase the protein unfolding starts off increasing at a low slope but then over
a narrow temp/salt conc the slope rapidly increases which suggests that there is
cooperativity in protein unfolding. unfolding at one part of the protein could destabilize
other parts of the protein making them more likely to unfold
o What did Sanger’s method and Edman’s degradation allow for,
o - Sanger’s method allowed the n-terminus amino acid residue to be determined. Edman’s
degradation allowed a sequence of amino acids to be determined. To determine the
sequence of a polypeptide, you can use Sanger’s method to determine the n-terminus
amino acids, then reduce the disulfide bonds to get linear peptides, digest these peptides
with different proteases, use Edman’s degradation to sequence each peptide fragment, and
then identify overlapping regions to piece the peptide back together and figure out their
sequences.
o how you would determine the sequence of a polypeptide containing one disulfide bond? 
o Once you figure out the sequences of both linear peptides, you just need to look at the
cysteines to figure out where the disulfide bonds are.