Isolating And Analyzing Proteins

Question 1

Which of the following is most critical for maintaining the naive tertiary structure of a globular protein?
A)hydrogen bonds between regions of the polypeptide chain in the folded protein
B)the hydrophobic effect of burying non-polar residues in the protein interior
C)disulfide bonds between Cys residues in the proteins
D)salt bridges (i.e. ionic bonds between opposite charged side chains).
E) none of the above

Answer 1

B)the hydrophobic effect of burying non-polar residues in the protein interior
All of these factors (plus optimized van der waals forces in the packed protein interior) can contribute to stabilize the native fold proteins structure, but the hydrophobic effect of burying non-polar amino acids is dominant for most proteins. The major factor opposing the native state is the increase in the chain entropy (i.e. conformational possibilities) in the unfolded state.

Question 2

Mutagenic replacement of isoleucine (Ile) with glycine (Gly) in the interior of a globular enzyme would most likely readily in results of loss of function because of ………
A)altered packing due to the smaller Glycine side chain
B)loss of disulfide bond formation by Ile
C)loss of the negative charge of Ile
D)the increased hydrophobic nature of Gly versus Ile
E)loss of hydrogen bonding from the Ile side chain

Answer 2

A)altered packing due to the smaller Glycine side chain
This would be an example of non- conservative amino acid replacement (i.e. by amino acid different in size, charge and/or chemical properties), and would not normally be tolerated in the interior of a tightly packed globular protein. Isoleucine is a fairly hydrophobic (non-polar) amino acid with a side chain thats is not ionizable, lacks a sulfhydryl group, and is incapable of forming hydrogen bonds.

Question 3

The 6X histadine affinity tag:
A)causes proteins to enter a stationary phase and elute from a column more slowly
B)is composed of a mixture of histadine and arginine residues
C)allows proteins to be purified by an ion exchange chromatography
D)binds tightly to cations such as Ni2+
E)permanently binds to proteins to a stationary phase

Answer 3

D)binds tightly to cations such as Ni2+

Question 4

During cation exchange chromatography:
A)positive charges on the surface of proteins interact with negatively charged resin.
B)negative charges on the surface of proteins interact with negatively charged resin
C)positive charged on the surfaces of proteins interact with positively charged resin
D)negative charges on the surface of proteins interact with positively charged resin
E)all of the above

Answer 4

A)positive charges on the surface of proteins interact with negatively charged resin.