PURIFICATION AND SUBSTRATE-SPECIFICITY OF AN ANGIOTENSIN-CONVERTING ELASTASE-2 FROM THE RAT MESENTERIC ARTERIAL BED PERFUSATE

A soluble angiotensin (Ang) II-generating enzyme has been purified to homogeneity from the rat mesenteric arterial bed (MAB) perfusate by a combination of gel filtration and affinity chromatographies. The enzym e is a glycoprotein of 28.5 kDa (SDS-PAGE), whose N-terminal sequence is identical with that of the rat pancreatic elastase-2; therefore the enzyme will henceforth be referred to as rat MAB elastase-2. When Ang I was used as the substrate, the enzyme specifically released Ang II and the dipeptide His-Leu (K-m = 36 mu M; K-cat = 1530 min(-1)). The c atalytic efficiency (K-cat/K-m = 42.5 min(-1) mu M-1) of this reaction was comparable to those of other known Ang I-converting enzymes. The proteolytic specificity of the purified enzyme toward mellitin, oxidiz ed insulin B chain, somatostatin-14 and renin substrate tetradecapepti de suggested that the enzyme-substrate interaction was defined by an e xtended substrate binding site, typical of elastases-2 of pancreatic o rigin. According to the sensitivity of the rat MAB elastase-2 to vario us inhibitors this enzyme could be described as a member of the chymos tatin-sensitive group of Ang II-forming serine proteases. The localiza tion and biochemical properties of this enzyme suggest that it might p lay a role in the regional control of vascular tonus. (C) 1998 Elsevie r Science B.V. All rights reserved.