Inhibition of convertase-related processing of proendothelin-1.

The biologically inactive precursor proendothelin-1 (proET-1) is initially processed intracellularly to the intermediate big endothelin peptide (big ET-1) before its conversion to the endothelin-1 (ET-1) peptide by the endothelin-converting enzyme (ECE). We recently demonstrated that purified furin, a calcium-dependent serine endoprotease belonging to the family of mammalian convertases, cleaved proET-1 in vitro and hence produced the physiologically relevant big ET-1 peptide. Therefore, furin becomes a candidate proET-1-cleaving enzyme responsible for the initial biosynthetic processing steps of this precursor. In this study we examined the inhibitory properties of two convertase inhibitors, i.e., the decanoyl-Arg-Val-Lys-Arg-chloromethylketone (dec-RVKR-cmk) and the alpha 1-antitrypsin Portland (AT-PDX) on proET-1 processing. When purified furin or PACE4, another mammalian convertase, was incubated in the presence of dec-RVKR-cmk, proET-1 processing was completely abolished. In the presence of purified AT-PDX, furin-related cleavage of proET-1 was abolished but not PACE4 processing. In addition, incubation of endothelial cells with dec-RVKR-cmk inhibited production of ET-1 but did not significantly alter the levels of the prostanoid PGI2. These results indicate that convertase-related processing of proET-1 can be inhibited in vitro and in vivo, and that convertase-specific processing may be prevented with AT-PDX.