THE T-TRANSITION TO R-TRANSITION IN THE COPPER(II)-SUBSTITUTED INSULIN HEXAMER - ANION COMPLEXES OF THE R-STATE SPECIES EXHIBITING TYPE-1 AND TYPE-2 SPECTRAL CHARACTERISTICS

The R-state conformation of the Cu(II)-substituted insulin hexamer has been identified, and a number of its derivatives have been studied via H-1 NMR, ESR, and UV-visible spectroscopy. This work establishes that the Cu(II)-substituted insulin hexamer undergoes an analogous T to R conformational transition in solution that has been identified previously for Zn(II)- and Co(II)-insulin hexamers [Roy, M., Brader, M. L., Lee, R. W.-K., Kaarsholm, N. C., Hansen, J., & Dunn, M. F. (1989) J. Biol. Chem. 264, 19081-190851. The data indicate that each Cu(II) center of the R-state Cu(II)-insulin hexamer possesses a coordination site that is accessible to anions from solution. Both phenol and anionic ligands that coordinate to the Cu(II) ions are required to generate the necessary heterotropic interactions that stabilize the R-state structure. With phenylmethylthiolate (PMT), a Cu(II)-R6 adduct that displays the spectral features of blue (type 1) copper proteins is obtained. This complex is proposed to embody a pseudotetrahedral Cu(II)N3S(PMT) chromophore, in which N is HisB10 (imidazolyl). The remaining ligands examined gave rise to Cu(II)-R6 adducts that possessed the spectral characteristics of normal (type 2) Cu(II) proteins. Under reducing conditions, Cu(I)-T6 and Cu(I)-R6 hexamers have been identified.