ACS BIOCHEMISTRY FINAL EXAM STUDY GUIDE | Actual Questions
and Answers Latest Updated 2024/2025 (Graded A+)
Henderson-Hasselbach Equation - ✔✔pH = pKa + log ([A-] / [HA])
Salting Out (Purification) - ✔✔Changes soluble protein to solid precipitate. Protein precipitates when
the charges on the protein match the charges in the solution.
Size-Exclusion Chromatography - ✔✔Separates sample based on size with smaller molecules eluting
later.
Ion-Exchange Chromatography - ✔✔Separates sample based on charge. CM attracts +, DEAE attracts -.
May have repulsion effect on like charges. Salt or acid used to remove stuck proteins.
Hydrophobic/Reverse Phase Chromatography - ✔✔Beads are coated with a carbon chain. Hydrophobic
proteins stick better. Elute with non-H-bonding solvent (acetonitrile).
Affinity Chromatography - ✔✔Attach a ligand that binds a protein to a bead. Elute with harsh chemicals
or similar ligand.
SDS-PAGE - ✔✔Uses SDS. Gel is made from cross-linked polyacrylamide. Separates based off of mass
with smaller molecules moving faster. Visualized with Coomassie blue.
SDS - ✔✔Sodium dodecyl sulfate. Unfolds proteins and gives them uniform negative charge.
Isoelectric Focusing - ✔✔Variation of gel electrophoresis where protein charge matters. Involves
electrodes and pH gradient. Protein stops at their pI when neutral.
Iodoacetate - ✔✔Adds carboxymethyl group on free -SH groups. Blocks disulfide bonding.
Homologs - ✔✔Shares 25% identity with another gene
,Orthologs - ✔✔Similar genes in different organisms
Ramachandran Plot - ✔✔Shows favorable phi-psi angle combinations. 3 main "wells" for α-helices, ß-
sheets, and left-handed α-helices.
α-helices - ✔✔Ala is common, Gly & Pro are not very common. Side-chain interactions every 3 or 4
residues. Turns once every 3.6 residues. Distance between backbones is 5.4Å.
Helix Dipole - ✔✔Formed from added dipole moments of all hydrogen bonds in an α-helix. N-terminus is
δ+ and C-terminus is δ-.
ß-sheet - ✔✔Either parallel or anti-parallel. Often twisted to increase strength.
Anti-parallel ß-sheet - ✔✔Alternating sheet directions (C & N-termini don't line-up). Has straight H-
bonds.
Parallel ß-sheet - ✔✔Same sheet directions (C & N-termini line up). Has angled H-bonds.
ß-turns - ✔✔Tight u-turns with specific phi-psi angles. Must have gly at position 3. Proline may also be at
ß-turn because it can have a cis-omega angle.
Loops - ✔✔Not highly structured. Not necessary highly flexible, but can occasionally move. Very variable
in sequence.
Circular Dichroism - ✔✔Uses UV light to measure 2° structure. Can be used to measure destabilization.
Disulfide-bonds - ✔✔Bonds between two -SH groups that form between 2° and 3° structure.
ß-mercaptoethanol - ✔✔Breaks disulfide bonds.
, α-keratin - ✔✔formed from 2 α-helices twisted around each other. "Coiled coil". Cross-linked by
disulfide bonds.
Collagen - ✔✔Repeating sequence of Gly-X-Pro. 3 stranded "coiled coil". Contains gly core.
Myoglobin 4° Structure - ✔✔Symmetric homodimer,
Hemoglobin 4° Structure - ✔✔Tetramer. Dimer of dimers. α2ß2 tetramer.
Mechanism of Denaturants - ✔✔Highly soluble, H-binding molecules. Stabilize protein backbone in
water. Allows denatured state to be stabilized.
Temperature Denaturation of Protein - ✔✔Midpoint of reaction is Tm.
Cooperative Protein Folding - ✔✔Folding transition is sharp. More reversible.
Folding Funnel - ✔✔Shows 3D version of 2D energy states. Lowest energy is stable protein. Rough
funnel is less cooperative.
Protein-Protein Interfaces - ✔✔"Core" and "fringe" of the interfaces. Core is more hydrophobic and is
on the inside when interfaced. Fringe is more hydrophilic.
π-π Ring Stacking - ✔✔Weird interaction where aromatic rings stack on each other in positive
interaction.
Fe Binding of O2 - ✔✔Fe2+ binds to O2 reversible. Fe3+ has an additional + charge and binds to O2
irreversibly. Fe3+ rusts in O2 rich environments.
ϴ-value in Binding - ✔✔ϴ = (bound / total)x100%
and Answers Latest Updated 2024/2025 (Graded A+)
Henderson-Hasselbach Equation - ✔✔pH = pKa + log ([A-] / [HA])
Salting Out (Purification) - ✔✔Changes soluble protein to solid precipitate. Protein precipitates when
the charges on the protein match the charges in the solution.
Size-Exclusion Chromatography - ✔✔Separates sample based on size with smaller molecules eluting
later.
Ion-Exchange Chromatography - ✔✔Separates sample based on charge. CM attracts +, DEAE attracts -.
May have repulsion effect on like charges. Salt or acid used to remove stuck proteins.
Hydrophobic/Reverse Phase Chromatography - ✔✔Beads are coated with a carbon chain. Hydrophobic
proteins stick better. Elute with non-H-bonding solvent (acetonitrile).
Affinity Chromatography - ✔✔Attach a ligand that binds a protein to a bead. Elute with harsh chemicals
or similar ligand.
SDS-PAGE - ✔✔Uses SDS. Gel is made from cross-linked polyacrylamide. Separates based off of mass
with smaller molecules moving faster. Visualized with Coomassie blue.
SDS - ✔✔Sodium dodecyl sulfate. Unfolds proteins and gives them uniform negative charge.
Isoelectric Focusing - ✔✔Variation of gel electrophoresis where protein charge matters. Involves
electrodes and pH gradient. Protein stops at their pI when neutral.
Iodoacetate - ✔✔Adds carboxymethyl group on free -SH groups. Blocks disulfide bonding.
Homologs - ✔✔Shares 25% identity with another gene
,Orthologs - ✔✔Similar genes in different organisms
Ramachandran Plot - ✔✔Shows favorable phi-psi angle combinations. 3 main "wells" for α-helices, ß-
sheets, and left-handed α-helices.
α-helices - ✔✔Ala is common, Gly & Pro are not very common. Side-chain interactions every 3 or 4
residues. Turns once every 3.6 residues. Distance between backbones is 5.4Å.
Helix Dipole - ✔✔Formed from added dipole moments of all hydrogen bonds in an α-helix. N-terminus is
δ+ and C-terminus is δ-.
ß-sheet - ✔✔Either parallel or anti-parallel. Often twisted to increase strength.
Anti-parallel ß-sheet - ✔✔Alternating sheet directions (C & N-termini don't line-up). Has straight H-
bonds.
Parallel ß-sheet - ✔✔Same sheet directions (C & N-termini line up). Has angled H-bonds.
ß-turns - ✔✔Tight u-turns with specific phi-psi angles. Must have gly at position 3. Proline may also be at
ß-turn because it can have a cis-omega angle.
Loops - ✔✔Not highly structured. Not necessary highly flexible, but can occasionally move. Very variable
in sequence.
Circular Dichroism - ✔✔Uses UV light to measure 2° structure. Can be used to measure destabilization.
Disulfide-bonds - ✔✔Bonds between two -SH groups that form between 2° and 3° structure.
ß-mercaptoethanol - ✔✔Breaks disulfide bonds.
, α-keratin - ✔✔formed from 2 α-helices twisted around each other. "Coiled coil". Cross-linked by
disulfide bonds.
Collagen - ✔✔Repeating sequence of Gly-X-Pro. 3 stranded "coiled coil". Contains gly core.
Myoglobin 4° Structure - ✔✔Symmetric homodimer,
Hemoglobin 4° Structure - ✔✔Tetramer. Dimer of dimers. α2ß2 tetramer.
Mechanism of Denaturants - ✔✔Highly soluble, H-binding molecules. Stabilize protein backbone in
water. Allows denatured state to be stabilized.
Temperature Denaturation of Protein - ✔✔Midpoint of reaction is Tm.
Cooperative Protein Folding - ✔✔Folding transition is sharp. More reversible.
Folding Funnel - ✔✔Shows 3D version of 2D energy states. Lowest energy is stable protein. Rough
funnel is less cooperative.
Protein-Protein Interfaces - ✔✔"Core" and "fringe" of the interfaces. Core is more hydrophobic and is
on the inside when interfaced. Fringe is more hydrophilic.
π-π Ring Stacking - ✔✔Weird interaction where aromatic rings stack on each other in positive
interaction.
Fe Binding of O2 - ✔✔Fe2+ binds to O2 reversible. Fe3+ has an additional + charge and binds to O2
irreversibly. Fe3+ rusts in O2 rich environments.
ϴ-value in Binding - ✔✔ϴ = (bound / total)x100%
