ACS BIOCHEMISTRY EXAM
Henderson-Hasselbach Equation - Answer-pH = pKa + log ([A-] / [HA])
FMOC Chemical Synthesis - Answer-Used in synthesis of a growing amino acid chain to a polystyrene
bead. FMOC is used as a protecting group on the N-terminus.
Salting Out (Purification) - Answer-Changes soluble protein to solid precipitate. Protein precipitates
when the charges on the protein match the charges in the solution.
Size-Exclusion Chromatography - Answer-Separates sample based on size with smaller molecules eluting
later.
Ion-Exchange Chromatography - Answer-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 - Answer-Beads are coated with a carbon chain.
Hydrophobic proteins stick better. Elute with non-H-bonding solvent (acetonitrile).
Affinity Chromatography - Answer-Attach a ligand that binds a protein to a bead. Elute with harsh
chemicals or similar ligand.
SDS-PAGE - Answer-Uses SDS. Gel is made from cross-linked polyacrylamide. Separates based off of
mass with smaller molecules moving faster. Visualized with Coomassie blue.
SDS - Answer-Sodium dodecyl sulfate. Unfolds proteins and gives them uniform negative charge.
,Isoelectric Focusing - Answer-Variation of gel electrophoresis where protein charge matters. Involves
electrodes and pH gradient. Protein stops at their pI when neutral.
FDNB (1-fluoro-2,3-dinitrobenzene) - Answer-FDNB reacts with the N-terminus of the protein to produce
a 2,4-dinitrophenol derivative that labels the first residue. Can repeat hydrolysis to determine sequential
amino acids.
DTT (dithiothreitol) - Answer-Reduces disulfide bonds.
Iodoacetate - Answer-Adds carboxymethyl group on free -SH groups. Blocks disulfide bonding.
Homologs - Answer-Shares 25% identity with another gene
Orthologs - Answer-Similar genes in different organisms
Paralogs - Answer-Similar "paired" genes in the same organism
Ramachandran Plot - Answer-Shows favorable phi-psi angle combinations. 3 main "wells" for α-helices,
ß-sheets, and left-handed α-helices.
Glycine Ramachandran Plot - Answer-Glycine can adopt more angles. (H's for R-group).
Proline Ramachandran Plot - Answer-Proline adopts fewer angles. Amino group is incorporated into a
ring.
α-helices - Answer-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 - Answer-Formed from added dipole moments of all hydrogen bonds in an α-helix. N-
terminus is δ+ and C-terminus is δ-.
ß-sheet - Answer-Either parallel or anti-parallel. Often twisted to increase strength.
Anti-parallel ß-sheet - Answer-Alternating sheet directions (C & N-termini don't line-up). Has straight H-
bonds.
Parallel ß-sheet - Answer-Same sheet directions (C & N-termini line up). Has angled H-bonds.
ß-turns - Answer-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 - Answer-Not highly structured. Not necessary highly flexible, but can occasionally move. Very
variable in sequence.
Circular Dichroism - Answer-Uses UV light to measure 2° structure. Can be used to measure
destabilization.
Disulfide-bonds - Answer-Bonds between two -SH groups that form between 2° and 3° structure.
ß-mercaptoethanol - Answer-Breaks disulfide bonds.
α-keratin - Answer-formed from 2 α-helices twisted around each other. "Coiled coil". Cross-linked by
disulfide bonds.
Collagen - Answer-Repeating sequence of Gly-X-Pro. 3 stranded "coiled coil". Contains gly core.
Henderson-Hasselbach Equation - Answer-pH = pKa + log ([A-] / [HA])
FMOC Chemical Synthesis - Answer-Used in synthesis of a growing amino acid chain to a polystyrene
bead. FMOC is used as a protecting group on the N-terminus.
Salting Out (Purification) - Answer-Changes soluble protein to solid precipitate. Protein precipitates
when the charges on the protein match the charges in the solution.
Size-Exclusion Chromatography - Answer-Separates sample based on size with smaller molecules eluting
later.
Ion-Exchange Chromatography - Answer-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 - Answer-Beads are coated with a carbon chain.
Hydrophobic proteins stick better. Elute with non-H-bonding solvent (acetonitrile).
Affinity Chromatography - Answer-Attach a ligand that binds a protein to a bead. Elute with harsh
chemicals or similar ligand.
SDS-PAGE - Answer-Uses SDS. Gel is made from cross-linked polyacrylamide. Separates based off of
mass with smaller molecules moving faster. Visualized with Coomassie blue.
SDS - Answer-Sodium dodecyl sulfate. Unfolds proteins and gives them uniform negative charge.
,Isoelectric Focusing - Answer-Variation of gel electrophoresis where protein charge matters. Involves
electrodes and pH gradient. Protein stops at their pI when neutral.
FDNB (1-fluoro-2,3-dinitrobenzene) - Answer-FDNB reacts with the N-terminus of the protein to produce
a 2,4-dinitrophenol derivative that labels the first residue. Can repeat hydrolysis to determine sequential
amino acids.
DTT (dithiothreitol) - Answer-Reduces disulfide bonds.
Iodoacetate - Answer-Adds carboxymethyl group on free -SH groups. Blocks disulfide bonding.
Homologs - Answer-Shares 25% identity with another gene
Orthologs - Answer-Similar genes in different organisms
Paralogs - Answer-Similar "paired" genes in the same organism
Ramachandran Plot - Answer-Shows favorable phi-psi angle combinations. 3 main "wells" for α-helices,
ß-sheets, and left-handed α-helices.
Glycine Ramachandran Plot - Answer-Glycine can adopt more angles. (H's for R-group).
Proline Ramachandran Plot - Answer-Proline adopts fewer angles. Amino group is incorporated into a
ring.
α-helices - Answer-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 - Answer-Formed from added dipole moments of all hydrogen bonds in an α-helix. N-
terminus is δ+ and C-terminus is δ-.
ß-sheet - Answer-Either parallel or anti-parallel. Often twisted to increase strength.
Anti-parallel ß-sheet - Answer-Alternating sheet directions (C & N-termini don't line-up). Has straight H-
bonds.
Parallel ß-sheet - Answer-Same sheet directions (C & N-termini line up). Has angled H-bonds.
ß-turns - Answer-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 - Answer-Not highly structured. Not necessary highly flexible, but can occasionally move. Very
variable in sequence.
Circular Dichroism - Answer-Uses UV light to measure 2° structure. Can be used to measure
destabilization.
Disulfide-bonds - Answer-Bonds between two -SH groups that form between 2° and 3° structure.
ß-mercaptoethanol - Answer-Breaks disulfide bonds.
α-keratin - Answer-formed from 2 α-helices twisted around each other. "Coiled coil". Cross-linked by
disulfide bonds.
Collagen - Answer-Repeating sequence of Gly-X-Pro. 3 stranded "coiled coil". Contains gly core.
