EFFECT OF SECONDARY STRUCTURE ON THE RATE OF DEAMIDATION OF SEVERAL GROWTH-HORMONE RELEASING-FACTOR ANALOGS

The objective of this study was to determine whether the rates of deam idation of Asn8 in selected growth hormone releasing factor (GRF) anal ogs were related to the peptide's secondary structures in solution. Bo vine or human [Leu27]GRF(I-32)NH2 (both having Gly at position 15), [A la15 Leu27]bGRF(I-32)NH2 and [Pro15 LeU27]bGRF(I-32)NH2 were used as m odel peptides. The peptide helical content (assessed by CD) increased with the increasing methanol concentration and was as follows: 7, 12 a nd 18% in 0% MeOH; 24, 48 and 52% in 40% MeOH; and 41, 77 and 81% in 8 0% MeOH for Pro15 LeU27 bGRF(I-32)NH2, [LeU27]hGRF(1-32)NH2 and Ala15 LeU27 bGRF(1-32)NH2, respectively. 2D NMR studies done in the presence of 40% CD3OH indicated more helical structure for the Ala15 analog as compared to [Leu27]hGRF(1-32)NH2. In both these peptides Asn8 was inc luded in the helical region. In contrast, the lack of conformational i nformation for the Pro15 analog indicated little helical structure aro und Asn8. The peptides' deamidation rates decreased and their half-liv es increased with increasing MeOH concentrations. At 40% MeOH, the lea st helical Pro15 bGRF analog (t1/2 = 10.78 h) deamidated 1.5 and 2 tim es faster than its Gly15 (t1/2 = 15.74 h) and Ala15 (t1/2 = 21.53 h) c ounterparts, respectively. This study indicates that helical environme nt around Asn8 in GRF makes this residue less prone to deamidation. (C ) Munksgaard 1993.