title: Synthesis of phospho-urodilatin by combination of global phosphorylation with the segment coupling approach.
abstract: The chemical synthesis of biologically active phosphorylated urodilatin (CDD/ANP-95-126) was achieved by using a strategy of coupling protected peptide segments in solution. Three protected peptide segments corresponding to urodilatin (1-14) with side chain-unprotected Ser10, (15-24) and (25-32) were prepared manually using Fmoc chemistry on an aminopropyl polystyrene resin with the super acid-labile HMPB linker. For the coupling of segments, the carboxy group of the C-terminal segment (25-32) was converted into the tert-butyl ester by treatment with TBTA. The protected peptide segments were coupled in the presence of EDC/HOOBt or TBTU/HOBt to yield fully protected urodilatin with a free hydroxy function at Ser10. Introduction of the phosphate was performed with Et2NP(OtBu)2 and tetrazole followed by oxidation of the phosphite. Alternatively, a prephosphorylated protected segment (1-14) was used in the segment condensation. Our investigations indicate that both pathways, phosphorylation of protected urodilatin in solution and use of a prephosphorylated building block, are suitable methods to obtain a large phosphopeptide of high purity without formation of H-phosphonates or other by-products.
authors.affiliation: Niedersächsisches Institut für Peptid-Forschung (IPF), Hannover, Germany.
authors.forename: H
authors.initials: H
authors.lastname: Mostafavi
authors.forename: S
authors.initials: S
authors.lastname: Austermann
authors.forename: W G
authors.initials: WG
authors.lastname: Forssmann
authors.forename: K
authors.initials: K
authors.lastname: Adermann
mesh_terms: D000595:Amino Acid Sequence; D009320:Atrial Natriuretic Factor / Q000138:chemical synthesis* / Q000737:chemistry; D002851:Chromatography, High Pressure Liquid; D006863:Hydrogen-Ion Concentration; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000138:chemical synthesis* / Q000737:chemistry; D010748:Phosphopeptides / Q000737:chemistry; D010766:Phosphorylation
chemical_list: D010446:Peptide Fragments; D010748:Phosphopeptides; C103122:phospho-urodilatin; D009320:Atrial Natriuretic Factor