Question

Peptides can be separated using an ion‑exchange column based on their isoelectric (pI) values. At which pH values would two different peptides, one with a pI of 5.3 and the other with a pI of 8.5 , bind to a cation‑ and anion‑exchange column? Each peptide may be capable of binding to each column at more than one pH value.

Answer 1

At pH 3.5:

Peptide A with pI 5.1 and peptide B with pI 8.5 will both bind to the anion-exchange column.

At pH 6.5:

Peptide A with pI 5.1 will bind to the cation-exchange column, while peptide B with pI 8.5 will bind to the anion-exchange column.

At pH 10.8:

Peptide A with pI 5.1 and peptide B with pI 8.5 will both bind to the cation-exchange column.

Step-by-step explanation:

Ion-exchange column chromatography is used for separating proteins and other molecules in a solution based on differences in net charge. Negatively charged molecules bind to positively charged solid supports called the anion-exchange column, whereas, positively charged molecules bind to negatively charged solid supports known as cation-exchange column.

A peptide can acquire a net charge if the pH of the medium is either above or below its isoelectric point,pI. At pH above pI, the peptide acquires a net negative charge and can bind to a positive support. At pH above pI, the peptide acquires a net positive charge and can bind to a negative support. The greater the difference between pH and pI, the tighter the peptide will bind to the column.

At pH 3.5:

Peptide A with pI 5.1 and peptide B with pI 8.5 will both bind to the anion-exchange column.

At pH 6.5:

Peptide A with pI 5.1 will bind to the cation-exchange column, while peptide B with pI 8.5 will bind to the anion-exchange column.

At pH 10.8:

Peptide A with pI 5.1 and peptide B with pI 8.5 will both bind to the cation-exchange column.