Separation of cis/trans conformers of human and salmon calcitonin by low temperature capillary electrophoresis.
Conformer-specific recognition of peptides and proteins has often been proved with the aid of indirect methods. Here we provide an analytical approach for a direct investigation of separated isomers. Cis/trans conformers of the peptide hormones human (hCT) and salmon (sCT) calcitonin exhibit different migration properties in capillary zone electrophoresis at subambient temperatures. Calcitonin consists of 32 amino acids with two proline residues incorporated. It is the longest unstructured peptide for which a conformer separation by capillary electrophoresis has yet been achieved. Lowering the temperature yielded a splitting into two and three peaks for sCT and hCT, respectively, in acidic buffer. The peak ratios of 66:34 for sCT and 71:23 for hCT are in good agreement with the conformer distribution previously reported from nuclear magnetic resonance (NMR) studies. The addition of different organic modifiers (5-20% v/v) to the running buffer does not improve the separation. The observed merging of conformer peaks in buffer containing 20% v/v 2,2,2-trifluoroethanol (TFE) is attributed to structure formation.
Thunecke
F
F
Max-Planck Research Unit Enzymology of Protein Folding, Halle/Saale, Germany.
Fischer
G
G
D000595:Amino Acid Sequence; D000818:Animals; D002116:Calcitonin / Q000737:chemistry / Q000302:isolation & purification*; D019075:Electrophoresis, Capillary / Q000379:methods*; D006801:Humans; D008968:Molecular Conformation; D008969:Molecular Sequence Data; D012474:Salmon; D017386:Sequence Homology, Amino Acid; D013696:Temperature
C028815:salmon calcitonin; D002116:Calcitonin