21] Occurrence, detection, and biosynthesis of carboxy-terminal amides

Abstract

Publisher Summary This chapter describes the occurrence, detection, and biosynthesis of carboxy-terminal amides. The existence of an amide rather than a free α -carboxyl group at the COOH-terminus of peptides is found in numerous peptides secreted by tissues of many different organisms such as gastrin, cholecystokinin, substance P, secretin, vasoactive intestinal peptide, and thyrotropin-releasing hormone. This modification has only been detected in rather small polypeptides, ranging in size from the tripeptide thyrotropin releasing hormone to scorpion toxin II, which contains 64 amino acids. Terminal amides are generally detected by the change in electrophoretic mobility caused by the modification of the carboxyl end. A more general method to search for amidated peptides is described by Tatemoto and Mutt. The crude or purified peptide sample is degraded with a protease of broad specificity, such as thermolysin, subtilisin, or elastase, and the total digest is subsequently treated with dansyl chloride to yield the dansylated peptides and amino acids. From alkaline solutions, the dansyl derivatives of amidated amino acids and peptides can be preferentially extracted into ethyl acetate. As peptides are generally derived from larger precursors, it is noticeable that the specificity for amidation must reside in the structure of these larger forms, most likely in the amino acids adjacent to the terminal residue of the end product.