Folding Paradox - Levinthal's paradox states that there are approximately 10⁵⁰ possible conformations for a protein, such as ribonuclease (124 residues). If one new conformation could be attempted every 10^-13 seconds, it would still take over 10³⁰ years to randomly test all of the possibilities, yet ribonuclease can completely fold in about a minute. Thus, folding must not be a completely random phenomenon.

Pathway Model - The "pathway" model of protein folding is depicted at the left. Nucleation is critical because it is much more difficult to begin an helix than to extend it. Nucleation may start at a number of points and all of these partially folded structures can be "funneled" by energy minimizations toward the final state (Figure 6.24). Thus, Levinthal's paradox is averted.

Notice that the funnel in Figure 6.24 has energetic "traps", which correspond to local free energy minima associated with incorrectly folded states. Fortunately, the cell has ways to assist incorrectly folded proteins to find the proper conformation.

Common Errors - One of the most common folding errors occurs via cis-trans isomerization of the amide bond adjacent to a proline residue (see here). Proline is the only amino acid in proteins that forms peptide bonds in which the trans isomer is only slightly favored (4 to 1 versus 1000 to 1 for other residues). Thus, during folding, there is a significant chance that the wrong proline isomer will form first. It appears that cells have enzymes to catalyze the cis-trans isomerization necessary to speed correct folding.

Disulfide Bond Formation - Proteins with disulfide bonds have a built-in advantage if they are denatured with their disulfide bonds intact. The intact disulfide bonds eliminate many degrees of freedom associated with denaturation, so fewer events need to occur to bring about the correctly folded state. This can be verified by removing the disulfide bonds of a protein and then denaturing it. Refolding of this polypeptide occurs, but at a slower rate than when the disulfides are left intact (see Figure 6.25). Interestingly, disulfide bonds not found in the native structure sometimes form during intermediate stages of folding. Also, the folding process can be aided by enzymes that make disulfide bonds.

Chaperonins - In addition to the enzymes mentioned previously that assist with proper folding (e.g., cis-trans isomerase for proline and disulfide bond making enzymes), cells have a class of proteins called chaperonins, which "chaperone" a protein to help keep it properly folded and non-aggregated. Aggregration is a problem for unfolded proteins because the hydrophobic residues, which normally are deep inside of a protein, may be exposed when the protein is released from the ribosome. If they are exposed to hydrophobic residues in other strands, the two strands may associate with each other hydrophobically (to aggregate) instead of folding properly. The GroEL-ES complex from E. coli is one such chaperone system. It provides a central cavity in which new protein chains can be "incubated" until they have folded properly (Figure 6.26).

1. The Chaperonin Home Page

2. Protein Folding