Explain the function of each reagent, ezyme, procedure and results at each of st

Question

Explain the function of each reagent, ezyme, procedure and results at each of steps 1-7.

For The amino acid composition following: (1). Treatment with DTT and (2) acid hydrolosys of 1 nanomole of the peptide gives the following results.

Ala .92nm

Ser .73nm

Lys .97nm

Phe .88nm

Val .99nm

Glu .81nm

Pro 1.01nm

Cys .92nm

What is your interpretation of this data and what did step 1 and 2 do?

Estimate the MW of the octapeptide to the nearest 100. ______Da (how did you get this number)

Seperate samples of the reduced peptide are subjected to the following treatments:

After treatment with dansyl chloride at ph 8 followed by acid hydrolysis, you obtain an odd colored derivative that is inconclusive. When you react a fresh sample with ninhydrin however, you obtain a nie brownish yellow derivative that you recognize is due to the amino acid ________. Explain.

After treatment with carboxypeptidase, you obtain free Ser, plus a little Val. What does enzyme do? What is your conclusion and why?

The peptide has weak UV absorbance at 240nm. Interpret.

Explanation / Answer

1)

The interpretation:

The DDT cleaves the disulphide bonds between the two cystine amino acids, so this protein is not a liner peptide. Acid hydrolysis of one nm peptide gives almost equimolar concertation of all amino acids and it yields 8 amino acids. Hence it is an octapeptide.

2)

The general molecular weight of an amino acid is 110 Da. It is an octapeptide, so 8×110 = 880, but during peptide bond formation for one peptide bone one water molecules is released. This octapeptide has 7 peptide bonds; therefore we need to deduct 7 water molecules molecular weight from 880-126 = 754 Da.

Thus, the molecular weight of peptide is 745 Da.

3)

The amino acid proline gives yellow color with ninhydrin, which is because proline has an imno-group in its structure.

4)

Carboxypeptidase cleaves the C-terminal amino acid of peptide, as in interpretation I said that this octapeptide is not liner, hence it has two C-terminal and 1 N-terminal amino acids. So, Ser is one of the C-terminal and Val is another C-terminal amino acid.

5)

The Peptide has weak UV absorbance at 240 nm; this is because proteins which contain aromatic amino acids have maximum absorption at 280 nm. Our octapeptide has one aromatic amino acid “Phe.” So, it has less peak at 240 nm.

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