Problem Set 4: Chapter 4: The three dimensional structure of proteins

1) Name four factors (bonds or other forces) that contribute to stabilizing the native structure of a protein, and describe one condition or reagent that interferes with each type of stabilizing force. Ans: Among forces that stabilize native protein structures are (a) disulfide bonds, (b) hydrogen bonds, (c) hydrophobic interactions, and (d) ionic interactions. Agents that interfere with these forces are (a) mercaptoethanol or dithiothreitol, (b) pH extremes, (c) detergents and urea, and (d) changes in pH or ionic strength, respectively. 2) Describe three of the important features of the -helical polypeptide structure predicted by Pauling and Corey. Provide one or two sentences for each feature. Ans: The -helical structure of a polypeptide is tightly wound around a long central axis; each turn of the right-handed helix contains 3.6 residues and stretches 5.4 along the axis. The peptide NH is hydrogen-bonded to the carbonyl oxygen of the fourth amino acid along the sequence toward the amino terminus. The R groups of the amino acid residues protrude outward from the helical backbone. 3) Each of the following reagents or conditions will denature a protein. For each, describe in one or two sentences what the reagent/condition does to destroy native protein structure. (a) (b) (c) (d) urea high temperature detergent low pH

Ans: (a) Urea acts primarily by disrupting hydrophobic interactions. (b) High temperature provides thermal energy greater than the strength of the weak interactions (hydrogen bonds, electrostatic interactions, hydrophobic interactions, and van der Waals forces, breaking these interactions. (c) Detergents bind to hydrophobic regions of the protein, preventing hydrophobic interactions among several hydrophobic patches on the native protein. (d) Low pH causes protonation of the side chains of Asp, Glu, and His, preventing electrostatic interactions. 4) Once a protein has been denatured, how can it be renatured? If renaturation does not occur, what might be the explanation? Ans: Because a protein may be denatured through the disruption of hydrogen bonds and hydrophobic interactions by salts or organic solvents, removal of those conditions will reestablish the original aqueous environment, often permitting the protein to fold once again into its native conformation. If the protein does not renature, it may be because the denaturing treatment removed a required prosthetic group, or because the normal folding pathway requires the presence of a polypeptide chain binding protein or molecular chaperone. The normal folding pathway could also be mediated by a larger polypeptide, which is then cleaved (e.g., insulin). Denatured insulin would not refold easily.