Working With Proteins

1. Match the following techniques with how they are used to study proteins.
A) nuclear magnetic resonance
B) mass spectroscopy
C) X-ray crystallography
D) circular dichroism spectroscopy
E) SDS PAGE

  • determine exact mass of proteins and the sequence of peptides
  • determine protein structure using diffraction patterns and a computer program
  • measure amounts of protein secondary structures
  • determine structure of a protein in solution
  • determine molar mass
  • determine the primary sequence of a protein

2. Using complete sentences, briefly describe how each of the following are used to study proteins:
A) nuclear magnetic resonance
B) mass spectroscopy
C) X-ray crystallography
D) circular dichroism spectroscopy
E) SDS PAGE

3. A protein was digested with an enzyme and the following three peptides were purified by anion exchange chromatography at pH 6. Use the pKa values on page 156 in our text.
KANW ELDCARD SF ENW
A) Which peptide eluted first?
B) Second?
C) Third?
D) Fourth?
E) What enzyme was used to cut the peptide?

4. The following peptide was determined to have a Tryptophan as the N-terminus amino acid. After treatment with diothiothreitol and iodoacetate, the peptide was then cleaved and the resulting peptides were separated by anion exchange chromatography at pH 7 at 37°C. For all of the peptides, use the pKa values on page 156 in our text

The fragments are:
YSLDSETDKC
CHICAGQ
WINKA
YIKESMETS

A) What is the order of elution from the anion exchange column at pH 7.0:
1st.

2nd

3rd.

B) What enzyme was used to cleave the peptide?
C) Can you determine the sequence of the original peptide? What is it? Or, what are the possibilities?
D) What other experiments could be done to help determine the sequence?

5. The following peptide was cleaved with chymotrypsin. What are the products?
CANKCWINWILDCARD

6. The following three peptides were purified by cation exchange chromatography at pH 5. Use the pKa values you learned from page 156 in our book.
DETRIDT ANGELS WASHINGTN
a. Which peptide eluted first?
b. Second?
c. Third?

7. List the reagents for the following reactions that involve a peptide with “n” amino acids (n = the number of amino acids in the peptide):
A) Peptide(n) —-> labelled amino acid + free amino acids

B) Peptide(n) —-> labelled amino acid + peptide (n-1)

8. Why should a protein be treated with dithiothreitol, followed by iodoacetate if there are more than 2 cysteines in the protein?

Unless otherwise stated, the content of this page is licensed under Creative Commons Attribution-NonCommercial-ShareAlike 3.0 License