Q.27 A recombinant protein is secreted extracellularly in soluble form by an E. coli
culture. Which of the following downstream processes is/are involved in the
purification of the extracellular secreted protein?
(A) Cell disruption
(B) Membrane ultrafiltration
(C) Solubilisation of inclusion bodies
(D) Liquid chromatography

The correct answers are (B) Membrane ultrafiltration and (D) Liquid chromatography, as the protein is already secreted into the culture medium in soluble form, bypassing cell lysis steps.

Why These Processes Apply

Extracellular secretion simplifies purification by avoiding intracellular extraction. First, cells are separated via centrifugation, leaving the protein in the supernatant. Membrane ultrafiltration then concentrates it by retaining the protein while removing small impurities like salts. Liquid chromatography follows for high-purity separation based on size, charge, or affinity.

Option Breakdown

• (A) Cell disruption: Not needed, since the soluble protein is already outside intact E. coli cells; disruption applies to intracellular proteins.

• (B) Membrane ultrafiltration: Essential for initial concentration and clarification of the culture supernatant by filtering out smaller molecules.

• (C) Solubilisation of inclusion bodies: Irrelevant here, as this targets insoluble aggregates inside cells, not soluble extracellular proteins.

• (D) Liquid chromatography: Key final step for polishing purity through techniques like ion-exchange or affinity columns.