pH dependent conformational changes in the T-and R-states of insulin in solution: Circular dichroic studies in the pH range of 6 to 10

• Published in: Biochemical and biophysical research communications Vol. 186; no. 2; pp. 1115 - 1120
• Main Authors: Kim, Yesook, Shields, James E.
• Format: Journal Article
• Language: English
• Published: San Diego, CA Elsevier Inc 31.07.1992; Elsevier
• Subjects: Biological and medical sciences; Circular Dichroism; Conformational dynamics in molecular biology; Fundamental and applied biological sciences. Psychology; Humans; Hydrogen-Ion Concentration; Insulin - analogs & derivatives; Insulin - chemistry; Kinetics; Magnetic Resonance Spectroscopy - methods; Molecular biophysics; Protein Conformation; Solutions; Spectrophotometry, Ultraviolet - methods; Human; Protein hormone; Pancreatic hormone; Circular dichroism spectrometry; pH; Environmental factor; In vitro; Insulin; Conformational transition
• ISSN: 0006-291X; 1090-2104
• DOI: 10.1016/0006-291X(92)90862-F

Zinc insulin hexamer has been shown to undergo a phenol-induced T 6 to R 6 conformational transition in solution. Our circular dichroic (CD) studies demonstrate that insulin undergoes pH-dependent conformational changes over the pH range of 6–10 in the T-state and in the R- state. In order to determine which specific amino acid residues may be responsible for these pH-dependent changes, a series of insulin analogs were utilized. In the T-state, the pH dependent CD changes monitored in the far UV region have a pK of 8.2 and appear to be related to the titration of the A1-Gly amino group. Using the near UV CD a second pH-dependent conformational change was detected with a pK of 7.5 in the T-state. 1H N.M.R. studies suggest that B5-His may be responsible for this conformational transition. In the presence of m-cresol (R-state), the pK value was found to be 6.9. During this titration, the increased ellipticity for the R-state is diminishing as pH decreases from pH 8 to 6, and no difference in ellipticity was observed at 255 nm between T- and R-states at pH 6. Therefore, this may be due to the transition from the R back to the T-state.