Applications of BOP reagent in solid phase peptide synthesis. III: Solid phase peptide synthesis with unprotected aliphatic and aromatic hydroxyamino acids using BOP reagent

The BOP reagent [benzotriazol-1-yl-oxy-tris-(dimethylamino)phosphonium hexafluorophosphate], which has been shown to be ideally suited for solid phase synthesis, has now been found to be useful for solid phase synthesis using a minimal side-chain protection scheme. This new application of the BOP reagent was exemplified by the successful synthesis of the CCK-7 analog, Ac-Tyr(SO3H)-Met-Gly-Trp-Met-Thr(SO3H)-Phe-NH2, using unprotected Boc(hydroxy)-amino acids [Boc-Thr-OH and Boc-Tyr-OH]. N-Terminal acetylation was achieved under mild conditions by using the BOP coupling reaction with acetic acid. This procedure provided the unprotected (Tyr27, Thr32)-peptide-resin which is ready for the required sulfation on the solid support without selective side-chain deprotection of Ty27 and Thr32. Solid phase sulfation was evaluated under a variety of conditions and it was determined that disulfation was optimal using pyridine acetyl sulfate (38 equiv.) in pyridine at 45° for 4 h. Shorter reaction times or milder conditions lead to the formation of the Thr32 monosulfated analog. Cleavage of the disulfated analog from the PAM-resin was achieved using liquid ammonia and the product was purified by preparative hplc and fully characterized. The advantages of the new procedure are compared with the reported synthesis of CCK-7.