Protein Purification Techniques

Vocabulary flashcards covering cell lysis methods, centrifugation, liquid chromatography, ion exchange matrices, gel filtration, and affinity tagging based on the lecture.

POI (Protein of Interest)

The specific target protein in a cell lysate or sample mixture that is isolated and purified from other unwanted components.

Osmotic Lysis

A cell lysis method that uses a hypotonic solution to cause water to rush into animal cells until they burst; it only works on animal cells because plant and bacterial cells possess protective cell walls.

Mechanical Disruption

The use of physical shearing forces, such as high-speed blenders or a French press, to break cells open and liberate target proteins.

French Press

A mechanical instrument that applies very high pressure to squeeze cell samples through cell-sized or smaller pores to lyse them.

Sonication

A cell lysis technique that employs ultrasound waves combined with glass beads to shear cell membranes open.

Lysozyme

An enzyme that lyses bacterial cells by specifically cleaving components of the bacterial cell wall.

Denaturation

The unfolding of a protein's structure, often caused by conditions such as heat, leading to a loss of biological activity.

Centrifugation

A separation method that isolates proteins based on molecular mass or weight, where components with larger mass sediment faster under centrifugal force.

Svedberg Unit ($S$)

A unit representing the sedimentation coefficient in centrifugation, where a larger $S$ value correlates with a larger molecular mass.

Mobile Phase

The liquid solvent in liquid chromatography that carries the dissolved sample mixture through the column matrix.

Stationary Phase

The porous solid matrix packed within a liquid chromatography column that interacts with sample components to cause separation based on physical or chemical properties.

Carboxymethyl (CM) Matrix

A negatively charged (anionic) column matrix that acts as a cation exchanger by adsorbing positively charged molecules.

Diethylaminoethyl (DEAE) Matrix

A positively charged (cationic) column matrix containing protonated amine groups that acts as an anion exchanger by adsorbing negatively charged molecules.

Desorption

The process of releasing adsorbed proteins from a stationary matrix, typically accomplished by increasing salt concentration gradients or altering the pH.

Isoelectric Point ($pI$)

The specific pH value at which a molecule, polypeptide, or protein carries a net electrical charge of zero.

Gel Filtration Chromatography

A separation method (also known as size exclusion chromatography) that separates proteins by molecular weight, where the largest molecules elute first and the smallest elute last.

Affinity Chromatography

A capture purification technique that isolates a target protein based on its specific chemical attraction to a ligand attached to the solid column matrix.

His Tagging

A genetic engineering procedure that appends a polyhistidine sequence to the N- or C-terminus of a protein, allowing it to selectively bind to nickel ($\text{Ni}^{2+}$) ligands on an affinity column.