Total synthesis, purification, and characterization of human [Phe(p-CH2SO 3Na)52, Nle32,53,56, Nal55]-CCK20-58, [Tyr52, Nle32,53,56, Nal55]-CCK-58, and [Phe(p-CH2SO3Na)52, Nle32,53,56, Nal55]-CCK-58.

Journal Article (Journal Article)

The synthesis of [Phe(p-CH2SO3Na)52, Nle32,53,56 Nal55]-CCK20-58, [Tyr52, Nle32,53,56, Nal55]-CCK-58 and of [Phe(p-CH2SO3Na)52, Nle32,53,56, Nal55]-CCK-58 using the (9-fluorenylmethyloxy)-carbonyl (Fmoc) strategy on a 2,4-DMBHA resin is described. The crude peptide preparations were extremely complex when analyzed by RP-HPLC, capillary zone electrophoresis (CZE), and ion-exchange chromatography (IE-FPLC). We found that the most effective strategy for purification included cation-exchange chromatography followed by a RP-HPLC desalting step. The highly purified peptides (purity greater than 90%) were characterized by RP-HPLC, size exclusion HPLC (SEC), IE-FPLC, CZE, mass spectrometry, amino acid analysis, and Edman sequence analysis (for [Tyr52, Nle32,53,56, Nal55]-CCK-58). The results demonstrate the applicability of the 2,4-DMBHA resin for Fmoc solid-phase synthesis of long peptides amides (58 residues in length in this case) as well as the efficacy of an FPLC/RP-HPLC approach for the purification of very long, heterogeneous crude peptides, allowing a true assessment of the biological properties of these analogs to be carried out. [Phe(p-CH2SO3Na)52, Nle32,53,56, Nal55]-CCK20-58 was less than 1% as potent as CCK-8 while [Tyr52, Nle32,53,56, Nal55]-CCK-58 and [Phe(p-CH2SO3Na)52, Nle32,53,56, Nal55]-CCK-58 were inactive at the doses tested (< 0.01%).

Full Text

Duke Authors

Cited Authors

  • Miranda, MT; Craig, AG; Miller, C; Liddle, RA; Rivier, JE

Published Date

  • October 1993

Published In

Volume / Issue

  • 12 / 5

Start / End Page

  • 533 - 544

PubMed ID

  • 7511387

Pubmed Central ID

  • 7511387

International Standard Serial Number (ISSN)

  • 0277-8033

Digital Object Identifier (DOI)

  • 10.1007/BF01025118


  • eng

Conference Location

  • United States