Amino acids can exist as L-isomers or D-isomers. However with respect to the conformational stability, all amino acids are usually found in the L-isomeric form.

ANFINSEN’S EXPERIMENT (Reported in 1973)

• OBJECTIVE:  To establish that amino acid sequence dictates protein folding.
• EXPERIMENTAL INSIGHTS:

The roles of compounds like Urea, Guanidine HCl, β-mercaptoethanol, etc. on the refolding of an enzyme Riobnuclease A was elucidated through the experimental study. 8M Urea and Guanidine HCl together act as denaturants and disrupt the non-covalent bonds present in the protein. β-mercaptoethanol on the other hand acts as a reducing agent and disrupts the disulfide bonds, and when applied in large excess converts the disulfides to sulfhydryls. This denatured and reduced form of Ribonuclease A lacks enzymatic activity.

When Urea and β-mercaptoethanol removed are by the dialysis, the denatured and reduced Ribonuclease A spontaneously regains its native conformation and enzymatic activity. The removal of β-mercaptoethanol alone, however did not render the Ribonuclease A active and still lacks its enzymatic activity. It was also observed that trace amounts of β-mercaptoethanol facilitated the correct disulfide bonding and thereby enhanced the formation of native Ribonuclease A.

• CONCLUSIONS:  The phenomenon of protein folding is governed by the amino acid sequence, which is the actual repository of all the genetic information.

Illustration: Anfinsen’s experiment