ACID STABILIZATION OF INSULIN

The effect of pH on the conformational stability of insulin was studie d. Surprisingly, the Gibbs free energy of unfolding increased approxim ately 30% by acidification. pH titration of insulin's conformational s tability is described by a transition involving a single proton with a n apparent pK(a) of 7.0. The acid stabilization of insulin's conformat ion was attributed to the protonation of histidine at position 5 on th e B-chain (H(B5)) as determined by H-1-NMR of the histidines, selectiv e amino acid alteration, and enthalpies of ionization. Further acidifi cation (at least to pH 2) does not decrease the free energy of unfoldi ng. A conformational change in the tertiary structure, as indicated by the near-UV circular dichroism spectrum, accompanies this change in s tability. We propose that this acid stabilization of insulin is physio logically important in maintaining insulin stability in the acid envir onment of the secretory/storage granules of the beta-cell of the pancr eatic islets of Langerhans.