# DBT JRF 2022 — Question 18

Exam: DBT JRF 2022 · Section: B · Topic: Biochemistry › Proteolysis & Trypsin Digestion · Difficulty: Medium

## Question

A protein with 10 lysines and 2 arginine residues was digested with trypsin and the peptide mixture was resolved on an SDS-PAGE. After silver staining, 10 bands were detected instead of 13. Select the plausible reasons from the following: A. Some of the Arginines and Lysines were inside the protein core. B. Some of the Arginines and Lysines were followed by a Proline residue. C. Some of the Arginines and Lysines were followed by a negatively charged residue. D. Some of the Lysines were modified at the epsilon Nitrogen atom. E. One of the Lysine or Arginine was at the extreme C-terminus.                                                                                                                                                                                                                                                                                               Choose the correct answer from the options given below:

## Options

- **A.** A, B, C and D.
- **B.** A, B, C and E.
- **C.** B, C, D and E.
- **D.** A, B, D and E.

## Answer

Option D

## Explanation

Trypsin digestion and missing bands on SDS-PAGE: Trypsin cleaves after Lys (K) or Arg (R), EXCEPT when followed by Proline (P). Given: 10 Lys + 2 Arg = 12 potential cleavage sites → 13 expected fragments. Observed: only 10 bands → 3 fewer cuts occurred. Possible reasons: A. Buried Lys/Arg: inaccessible to trypsin in native protein — but in SDS-PAGE, the protein is fully denatured; however, the DIGESTION was done on the native protein potentially, or it's a distracting option. Actually, if buried, trypsin cannot access them. B. Arg/Lys before Pro: trypsin does NOT cleave Lys-Pro or Arg-Pro bonds — well-established rule ✔ C. Lys/Arg before negatively charged residue: trypsin is somewhat less efficient but generally still cleaves — this is NOT a primary reason for missed cleavages. D. Modified Lys ε-N (e.g., acetylation or methylation of the ε-amino group): removes the positive charge, making the site unrecognised by trypsin ✔ E. C-terminal Lys or Arg: produces no extra fragment (the last residue at the C-terminus doesn't create an additional cut) — this reduces expected count by 1 ✔ Most valid reasons: B, D, E (Option 4 = A, B, D and E). Answer: A, B, D and E (Option 4).

Source: https://mypyqbuddy.com/questions/a-protein-with-10-lysines-and-2-arginine-6a104cd5920734408bc0c721