Structural studies on des-pentapeptide (B26-30)-insulin. III. State of aggregation, tyrosyl environment, and peptide backbone conformation in solution.
The three-dimensional structure of des-pentapeptide (B26--30)-insulin (DPI) in solution has been studied by several methods in view of the obvious fact that such knowledge would have much to bear on an understanding of the structure of the receptor-binding site of insulin. (1) The molecular weight of DPI has been studied by the gel filtration method. The results demonstrate that DPI exhibits no self-association behaviour but exists as monomers under the experimental conditions. This enables us to deduce that insulin binds its receptor in the monomeric form; its functional unit is the monomer. (2) The pH difference spectra, denaturation difference spectra, and temperature effect curves have been examined. The results indicate that all three tyrosyl side chains of DPI are unmasked and situated on the surface of the molecule. (3) The UV CD curves of DPI have also been studied. The far UV CD curve of DPI at pH 7.0 is similar to that of insulin, except that the negative value for [iota]220 decreases. This probably demonstrates that the peptide backbone conformation of DPI is very similar to that of insulin, except that it is slightly extended for the former. The near UV CD curve for DPI differs from that for insulin and has an extremum at 266nm as well as a low negative value of [iota]. The assignment of this extremum to Phe and/or -S-S- has been discussed. Taking into consideration the results given in the following papers, it may be concluded that the results of conformation research obtained support our previous working hypothesis that the hydrophobic surface in between the two monomers making up dimeric insulin constitutes a part of the receptor-binding site of insulin.
Structural studies on des-pentapeptide (B26-30)-insulin. III. State of aggregation, tyrosyl environment, and peptide backbone conformation in solution. The three-dimensional structure of des-pentapeptide (B26--30)-insulin (DPI) in solution has been studied by several methods in view of the obvious fact that such knowledge would have much to bear on an understanding of the structure of the receptor-binding site of insulin. (1) The molecular weight of DPI has been studied by the gel filtration method. The results demonstrate that DPI exhibits no self-association behaviour but exists as monomers under the experimental conditions. This enables us to deduce that insulin binds its receptor in the monomeric form; its functional unit is the monomer. (2) The pH difference spectra, denaturation difference spectra, and temperature effect curves have been examined. The results indicate that all three tyrosyl side chains of DPI are unmasked and situated on the surface of the molecule. (3) The UV CD curves of DPI have also been studied. The far UV CD curve of DPI at pH 7.0 is similar to that of insulin, except that the negative value for [iota]220 decreases. This probably demonstrates that the peptide backbone conformation of DPI is very similar to that of insulin, except that it is slightly extended for the former. The near UV CD curve for DPI differs from that for insulin and has an extremum at 266nm as well as a low negative value of [iota]. The assignment of this extremum to Phe and/or -S-S- has been discussed. Taking into consideration the results given in the following papers, it may be concluded that the results of conformation research obtained support our previous working hypothesis that the hydrophobic surface in between the two monomers making up dimeric insulin constitutes a part of the receptor-binding site of insulin.