title: Characterization of "thyroliberin-deamidating enzyme" as a post-proline-cleaving enzyme. Partial purification and enzyme-chemical analysis of the enzyme from anterior pituitary tissue.
abstract: An enzyme which catalyzes the deamidation of thyroliberin (TRF; less than Glu-His-Pro-NH2) has been purified 110-fold from extracts of bovine anterior pituitary by ammonium sulfate fractionation, ion exchange chromatography on DEAE-cellulose, and gel filtration. This enzyme of 76,000 molecular weight (as estimated by gel filtration) exhibits maximal activity at neutral pH (optimum pH 7.4 to 7.6) in buffers of high ionic strength supplemented with thiol-protecting agents. As indicated by the strong inhibition of the enzymatic activity by N-ethylmaleimide and Hg2+, as well as by the extreme sensitivity toward diisopropyl fluorophosphate, -SH, and -OH residues apparently represent essential functional groups of the enzyme. The stereospecific deamidation of TRF (Km = 4.1 . 10(-4) M) is inhibited competitively by TRF analogues which contain proline or by the proline containing biologically active peptides luliberin (LH-RF), oxytocin, vasopressin, angiotensin II, and Substance P. TRF analogues without proline or peptide amides without proline are ineffective. This enzyme cleaves the appropriate Pro-X bonds in luliberin, angiotensin II, pyroGlu-His-Pro-Gly-NH2, and the collagenase substrate Z-Gly-Pro-Leu-Gly-Pro. Thus, it may be characterized as a post-proline-cleaving enzyme.
mesh_terms: D000581:Amidohydrolases / Q000302:isolation & purification; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001667:Binding, Competitive; D002417:Cattle; D010450:Endopeptidases / Q000378:metabolism*; D007700:Kinetics; D010455:Peptides / Q000494:pharmacology; D010903:Pituitary Gland, Anterior / Q000201:enzymology*; D013379:Substrate Specificity; D013973:Thyrotropin-Releasing Hormone
chemical_list: D000596:Amino Acids; D010455:Peptides; D013973:Thyrotropin-Releasing Hormone; D010450:Endopeptidases; D000581:Amidohydrolases