Application of a peptide bank from porcine brain in isolation of regulatory peptides

Over the past years, the introduction of biological assay systems, random p eptide sequencing and orphan receptor screening has led to the isolation an d identification of new regulatory peptides with potential clinical impact. We have developed a method for separating peptides into about 300 fraction s from large amounts of porcine brain tissue. The preparation of this pepti de bank consists of three steps including ultrafiltration followed by catio n-exchange separation and reversed-phase chromatography. These fractions re present the peptide bank with desalted and lyophilized peptides from brain tissue. Molecular masses of the peptides in the fractions are determined by matrix-assisted laser desorption ionization MS and a mass data bank is sub sequently generated. For systematic analysis of the peptides, a subsequent two-step purification procedure is followed by Edman sequencing resulting i n the identification of different peptides. A survival assay with a neurona l cell line revealing the stimulatory and inhibitory activities is applied as a model to test the 300 fractions. This primary screen indicates that th e biological activities of the extracted peptides are easily characterized and, moreover, can be related to the biochemical entities. We conclude that the established peptide bank is an efficient and useful tool for the isola tion of regulatory brain peptides applying different purification strategie s. (C) 1999 Elsevier Science B.V. All rights reserved.