A protein biochemist attempted to determine the amino acid sequence of a decapep

Question

A protein biochemist attempted to determine the amino acid sequence of a decapeptide. Use the results fromt he trypsinm chymotrypsin, and cyanogen bromide treatments to suggest the amino acid sequence.

Trypsin digestion gave two fragments

T1: Ala, Arg, Phe, Pro, Thr, Trp, Tyr

T2: Lys, Met, Val

Chymotrpysin digestion gave three fragments with the following residues

CT1: Ala, Arg, Phe, Pro

CT2: thr, Trp

CT3: Lys, Met, Tyr, Val

N terminal analysis of the CT1 tetrapeptide indicated that the N-terminal amino acid was Ala. The cyanogen bromide gave only one single amino acid, methionine, and a nonapeptide. wWhat is a possible sequence of the decapeptide? Use three letter abbreviations

Explanation / Answer

The decapeptide is Met-Val-Lys-tyr-thr-trp-ala-arg-pro-phe

1. Cyanogenbromide-cleaves carbonyl side of Met (met and nonapeptide)

2. Trypsin cleaves on the C side of Lys and Arg but not on the N side of Pro.This means that Lys is the last aminoacid in T2 (only trypsin fragment containing Met).

Hence, Met-Val-lys-........

3. The middle portion of the sequence can be determined from the chymotrypsin digestion. Chymotrypsin cleaves on the C side of phe, trp, tyr but not on the N side of pro.

4. CT3 has Lys, met, tyr, val. Since met-val and tyr are first 3 aminoacids so tyr must be in 4th. This gives Met-Val-Lys-Tyr-....

5. CT1 has ala, arg, phe, pro, since CT cleaves C side of phe the last aminoacid should be Phe. Also CT1 has first aminoacid as ala. Moreover trypsin doesnot cleaves on the N side of pro. so the sequence of CT1 stands: ala-arg-pro-phe.

6. In CT2 chymotrypsin cleavages on the C side of trp. So the sequence of CT2 stands as thr-trp and is present in the middle.

Therefore the whole sequence of decapeptide stands as:

Met-Val-Lys-tyr-thr-trp-ala-arg-pro-phe

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