Abstract: | The trypsin-catalyzed coupling of bovine (Boc)2-desoctapeptide (B23-B30)-insulin with synthetic octapeptides, H-Gly-X2-X3-X4-Thr-Pro-Lys(Boc)-Thr-OH (X2 = Phe or Ala, X3 = Phe or Ala, X4 = Tyr or Ala), followed by deprotection and purification produced the AlaB24, ThrB30]-, AlaB25, ThrB30]-, and AlaB26, ThrB30]-analogs of bovine insulin in yields of 32, 35, and 32%, respectively. The biological activity of these analogs decreased in the order, normal insulin (ThrB30]-bovine insulin) = AlaB26-insulin > AlaB25-insulin > AlaB24-insulin, as assayed for receptor binding and some other biological effects, in contrast with the corresponding Leu-analogs of human insulin, in which the activity decreased in the order, normal insulin > LeuB24-insulin > LeuB25-insulin. The affinity to insulin antibodies greatly diminished in both AlaB24-insulin and LeuB24-insulin but not in the B25-substituted analogs. The CD spectra of the Leu- and the Ala-analogs were compared with those of normal insulins to show that no apparent correlation seems to exist between the decrease in biological activity and the conformational changes observed in solution. The effects of organic solvents on the peptide-bond equilibrium and on the stability of trypsin are also discussed. |