Big endothelin‐1 structure important for specific processing by endothelin‐converting enzyme of bovine endothelial cells

Abstract

Phosphoramidon-sensitive endothelin-converting enzyme of bovine endothelial cells showed substrate selectivity for big endothelin-1 (ET-1) when compared to big ET-1(1-38), big ET-2(1-37), big ET-2(1-38) and big ET-3(1-41). To investigate the big ET-1 structure important for specific conversion by the endothelin-converting enzyme, we synthesized a series of truncated analogues of big ET-1, measured the hydrolysis of their Trp21-Val22 bonds, and found that a 16-residue peptide, big ET-1(19-34), is the minimal peptide sequence. This suggests that an unusually long carboxy-terminal sequence is required for big ET-1 conversion. Alanine substitution for individual amino acids in the carboxy-terminal region of big ET-1(19-34) demonstrated that His27, Val29, Pro30, Tyr31, Gly32, Leu33 and Gly34 are more important than Asn23, Thr24, Pro25, Glu26 and Val28 for eliciting efficient hydrolysis of the Trp21-Val22 bond, even though the former residues are located at more distant positions from the cleavage sites than are the latter. These results, together with the fact that big ET-2 and big ET-3 show heterogeneity in the big ET-1 residues His27, Val28, Val29 and Gly34, suggest that the His27-Val-Val-Pro-Tyr-Gly-Leu-Gly34 sequence in the carboxy-terminal region of big ET-1 plays the most important role in selective conversion by endothelin converting enzyme.