Degradation of bradykinin and its metabolites by rat brain synaptic membranes.
Bradykinin (BK) (Arg1-Pro2-Pro3-Gly4-Phe5-Ser6-Pro7-Phe8-Arg9) was degraded by rat brain synaptic membranes at a rate comparable to that found for Met-enkephalin, but approximately 40 times the rate for vasopressin and oxytocin. The catabolic pathway for BK and its metabolites was elucidated through the use of high performance liquid chromatography for metabolite identification and peptidase inhibitors for blocking specific cleavage sites. BK was hydrolyzed at three sites: at the -Phe5-Ser6- bond by metalloendopeptidase 24.15, at the -Pro7-Phe8- bond by an apparently novel peptidyl dipeptidase, and at the -Phe8-Arg9 bond by a carboxypeptidase B-like enzyme. Each enzyme contributed about equally to BK degradation under the assay conditions used. Some of the resulting metabolites were further hydrolyzed: BK(1-8) to BK(1-7) + Phe by a DFP inhibitable prolyl carboxypeptidase-like enzyme, BK(1-8) to BK(1-5) + BK(6-8) by metalloendopeptidase 24.15, BK(1-7) slowly to BK(1-5) by a second peptidyl dipeptidase which was captopril inhibited, and Phe-Arg to Phe + Arg by a bestatin-inhibited dipeptidase. A number of properties of the individual enzymes were determined including sensitivity to a variety of peptidase inhibitors. These results provide a starting point for investigating the potential physiological role of each enzyme in BK function in the brain.
Orawski
A T
AT
Department of Biochemistry and Biophysics, Loyola University of Chicago Stritch School of Medicine, Maywood, IL 60153.
Simmons
W H
WH
D000595:Amino Acid Sequence; D000818:Animals; D001920:Bradykinin / Q000378:metabolism*; D001921:Brain / Q000201:enzymology / Q000378:metabolism*; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D006863:Hydrogen-Ion Concentration; D008623:Mercaptoethanol / Q000494:pharmacology; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000032:analysis; D051381:Rats; D011919:Rats, Inbred Strains; D012995:Solubility; D013570:Synaptic Membranes / Q000201:enzymology / Q000378:metabolism*
D010446:Peptide Fragments; D008623:Mercaptoethanol; D001920:Bradykinin