MULTIPLE CONFORMATIONS AND PROLINE CIS-TRANS ISOMERIZATION IN SALMON-CALCITONIN - A COMBINED NUCLEAR-MAGNETIC-RESONANCE, DISTANCE GEOMETRY, AND MOLECULAR MECHANICS STUDY

The relationship between multiple conformations and proline cis-trans isomerization in salmon calcitonin (sCT) lias been investigated by H-1-NMR, distance geometry, and molecular mechanics. In different solvents, the isomerization of Pro(23) induces resonance heterogeneity for amino acids adjacent to it. NOESY experiments have related such heterogeneity to two different isoforms of the hormone, which interconvert into each other in polar solvents as well as in SDS micelles. The cis-trans ratio was found to depend upon the structure of the hormone. In water, the random-coil sCT showed a 35% cis population, while 16% cis was observed in the folded SDS-bound hormone. Such a decrease contributes, per se, a Gibbs free energy stabilization of 3.10 kJ mol(-1). Except for the 21-25 region, a common NOE pathway was found for both isomers. This is in agreement with the common secondary structure for both isoforms: a central helix and an extended C-terminal segment interacting with it. Calculations indicated that while the trans isomer is helical in the Thr(6)-Tyr(22) region, a shorter helix (Thr(6)-Lys(18)) is present in the cis isoform. It is concluded that isomerization of Pro(23) does not alter the three-dimensional structure of the hormone, although trans structures show a lower average NOE root mean square deviation and a higher relative stability. Both cis and trans structures satisfactorily reproduce the experimental data, although they do not fulfill the whole set of NOE restraints, pointing out the danger involved in structure calculation whenever the cis-trans equilibrium does not affect the global fold.