Protein Folding, Purification, Structure Determination, and Regulation

Vocabulary flashcards generated from lecture notes covering amino acid chemistry, thermodynamics of protein folding, levels of protein structure, purification methods, structural determination techniques, and post-translational regulatory mechanisms.

Amino Acid

The basic monomeric building block of proteins, consisting of an $\alpha$-carbon atom bonded to an amino group, a carboxyl group, a hydrogen atom, and a side chain ($R$).

L-amino acids

The specific optical stereoisomer (mirror-image form) of amino acids found exclusively in functional proteins.

Water-Water Hydrogen Bond

A non-covalent interaction defined geometrically by distance $d < 3.5\,\text{\AA}$ and angle $\theta < 30^\circ$, with a strength of $\sim 5\text{--}10\,\text{kcal/mol}$.

Hydrophilic Solute

A polar solute that dissolves in water because its formation of solute-water hydrogen bonds compensates for the energetic loss of disrupted water-water interactions.

Hydrophobic Solute

A non-polar solute that fails to dissolve in water because it cannot compensate for the energetic cost of disrupting water-water interactions.

Dehydration Reaction

A condensation reaction during which water is eliminated to form covalent bonds connecting amino acid monomers into polymers.

Peptide Bond

The covalent chemical bond formed between the carboxyl group of one amino acid and the amino group of another during a dehydration reaction.

Free Energy ($G$)

The thermodynamic quantity defined as $G = H - TS$, representing the total amount of energy available to perform useful work.

Enthalpy ($H$)

A measure of system heat defined as $H = U + PV$, where $U$ is internal energy (average interaction energy) and $PV$ is pressure-volume energy.

Entropy ($S$)

A thermodynamic state variable that measures system disorder and multiplicity (the number of available microstates or conformations).

van der Waals Force

An electrodynamic non-covalent force occurring between two fluctuating dipoles.

Salt Bridge

An electrostatic interaction formed between positively and negatively charged amino acid side chains (acids and bases).

Primary Structure

The specific linear sequence of amino acids joined together by peptide bonds in a polypeptide chain.

Secondary Structure

Local backbone conformations, such as right-handed $\alpha$-helices or $\beta$-sheets, stabilized by hydrogen bonding between backbone amide and carbonyl groups.

Ramachandran Plot

A plot of backbone dihedral torsional angles (phi and psi) displaying energetically allowed and disfavored secondary structure conformations.

Glycine

An amino acid lacking a side chain that exhibits high dihedral flexibility and acts as a secondary structure breaker.

Proline

An amino acid lacking a backbone amide hydrogen with constrained side chain and backbone angles, acting as a secondary structure breaker.

Tertiary Structure

The three-dimensional conformation of a single polypeptide chain, stabilized by electrostatics, van der Waals interactions, hydrogen bonds, and cysteine disulfide bridges.

Quaternary Structure

The complete three-dimensional complex formed by the assembly of multiple polypeptide subunits (e.g., hemoglobin).

Protein Domain

A modular region of a polypeptide chain that folds independently and self-stabilizes, usually corresponding to a specific cellular function.

Intrinsically Disordered Proteins

Proteins containing large, unstructured polypeptide regions that exist as random coils in cytosol and adopt distinct shapes upon binding specific partners.

Urea

A potent protein denaturant used at $8\,\text{M}$ concentration that destabilizes folded states by interacting strongly with both backbone atoms and non-polar groups.

Sodium Dodecyl Sulphate (SDS)

An anionic detergent that denatures proteins by coating hydrophobic residues, conferring a uniform negative charge-to-mass ratio.

Chaperones

Biological helper proteins (such as heat shock proteins or HSPs) that sequester unfolded polypeptide chains to facilitate proper folding.

Homogenization

A set of gentle mechanical procedures (ultrasound, detergents, high pressure, or rotating plungers) used during cell lysis to rupture cell plasma membranes.

Differential Centrifugation

A fractionation procedure that uses sequential spins at increasing rotational speeds to separate cell components based on size and density into pellets and supernatants.

Native PAGE

Polyacrylamide gel electrophoresis that separates non-denatured proteins based on both their native three-dimensional shape and net charge.

SDS-PAGE

Denaturing polyacrylamide gel electrophoresis that separates proteins strictly according to molecular length or mass due to uniform SDS binding.

Isoelectric Focusing PAGE

Electrophoresis technique that migrates proteins along a pH gradient until reaching their isoelectric point ($pI$), where net charge becomes zero.

2-D PAGE

A high-resolution protein separation technique combining Isoelectric Focusing (first dimension, based on $pI$) with SDS-PAGE (second dimension, based on mass).

X-ray Diffraction

A structural determination method that measures X-ray scattering by electron clouds in crystalline samples to map atomic arrangements.

NMR Spectroscopy

A structural method exploiting intrinsic nuclear magnetization to determine dynamic structures of small ($< 50\,\text{kD}$) soluble proteins in solution.

Cryo-EM

An imaging technique that combines thousands of 2D electron microscope projections of un-crystallized biomolecules into a high-resolution 3D structure.

Dissociation Constant ($K_d$)

The equilibrium ratio $\frac{k_{\text{off}}}{k_{\text{on}}}$, where lower numerical values represent higher protein-ligand binding affinity.

Phosphorylation

A post-translational modification adding a phosphate group to Ser, Thr, or Tyr residues, introducing a negative charge that regulates protein activity.

Palmitoylation

A covalent post-translational modification attaching a palmityl group to a Cys residue, anchoring the protein to lipid membranes.

Ubiquitination

The covalent attachment of ubiquitin molecules to substrate Lys residues, where K48-linked polyubiquitination signals for proteasomal degradation.

Allosteric Regulation

Control of protein function occurring when a regulatory ligand binds to a distinct regulatory site distant from the catalytic active site, driving conformational changes.