A scientist wants to separate three protein molecules by ion-exchange chromatography. The pH of the mobile phase of the column is maintained in such a way that the protein molecule (A) has a net charge of -2, protein molecule (B) has a net charge of +2 and protein molecule (C) has a net charge of +1. Which one of the molecule/s will elute first from a cation-exchange resin?
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Remember: "Cation-exchange" means it {exchanges} (and thus binds) cations. Anions (negatively charged molecules) will not bind at all and will be found in the flow-through, eluting first.
Step 1: Understanding the Concept:
Ion-exchange chromatography separates molecules based on their net charge.
Cation-exchange resin: The resin is negatively charged and binds positively charged molecules (cations).
Anion-exchange resin: The resin is positively charged and binds negatively charged molecules (anions).
Molecules with the same charge as the resin are repelled and elute first. Step 2: Detailed Explanation:
The resin is a cation-exchange resin, meaning its stationary phase is negatively charged.
The protein charges are: A (-2), B (+2), and C (+1).
Protein A (-2): Having a negative charge, it will be repelled by the negative resin. It will not bind and will pass through the column immediately, thus eluting first.
Proteins B (+2) and C (+1): Having positive charges, they will bind to the negative resin. They will be eluted later, typically by increasing the salt concentration of the mobile phase. Protein B (+2) will bind more tightly than Protein C (+1).
Step 3: Final Answer:
Protein molecule A, with a net negative charge, will be repelled by the cation-exchange resin and will elute first.
