Mammalian natriuretic peptides from atrial tissue may have a physiological role in maintaining water and electrolyte balance and vascular tone. The peptides that have been sequenced all contain the 21 amino acid sequence of the smallest active peptide, atriopeptin I. Since the primary gene product deduced from cDNA contains 152 amino acids, it is apparent that the natriuretic peptides arise through the action of a number of selective processing enzymes. We have already found an enzyme (atrial dipeptidyl carboxy hydrolase) that probably converts atriopeptin II to atriopeptin I. We will continue our research with this enzyme. Our research will also center on identifying and purifying other processing enzymes and studying their kinetic properties and substrate specificities as well as their molecular properties. This research will involve the use of synthetic substrates for delineating the specificity of the enzymes and synthetic inhibitors for distinguishing between them and other enzymes. Such inhibitors will be useful for purifying the enzymes by affinity chromatography and may be of pharmacological interest. We will also develop convenient methods of assaying these enzymes. We will purify the precursor to the active peptides, atriopeptigen, using thiol-disulfide interchange chromatography and gel permeation liquid chromatography. We will radiolabel atriopeptigen at its carbohydrate residues (if present) and/or at its reduced sulfhydryl groups. We will use the radiolabeled protein as a potential substrate for the processing enzymes we hope to identify. The cleavage pattern of atriopeptigen will be determined.
