Do the Structures of Big ET-1 and Big ET-3 Adopt a Similar Overall Fold? Consequences for Endothelin Converting Enzyme Specificity

• Published in: Biochemistry (Easton) Vol. 38; no. 6; pp. 1721 - 1726
• Main Authors: Cronin, N. B, Wallace, B. A
• Format: Journal Article
• Language: English
• Published: United States American Chemical Society 09.02.1999
• Subjects: Amino Acid Sequence; Animals; Aspartic Acid Endopeptidases - chemistry; Cattle; Circular Dichroism; Endothelin-1; Endothelin-3; Endothelin-Converting Enzymes; Endothelins - chemistry; Guinea Pigs; Humans; Hydrogen-Ion Concentration; Metalloendopeptidases; Mice; Models, Molecular; Molecular Sequence Data; Protein Conformation; Protein Folding; Protein Precursors - chemistry; Protein Structure, Secondary; Rabbits; Rats; Sequence Homology, Amino Acid; Substrate Specificity; Temperature
• ISSN: 0006-2960; 1520-4995
• DOI: 10.1021/bi981689b

Big ET-1 and big ET-3 are precursor peptides which render endothelin-1 (ET-1) and endothelin-3 (ET-3) relatively unreactive and resistant to proteolytic cleavage. Big ET-1 is cleaved in vivo by ECE-1 (endothelin-converting enzyme), and big ET-3 is also cleaved but apparently to a significantly lesser extent by this enzyme. To shed light on the relation between structure and function, circular dichroism (CD) spectroscopy and homology modeling were used to determine whether big ET-1 and big ET-3 adopt similar secondary and tertiary structures. Analyses of the CD spectra and thermal denaturation indicate they have similar secondary structures and thermal stabilities. Superposition of the modeled coordinates of both peptides indicates that they can adopt the same overall fold except in the C-terminal residues, 34−38 in big ET-1 and 34−41 in big ET-3. This region corresponds to an area of complete sequence heterogeneity between the two peptides. A model has been developed which has a loop for residues 27−30 (HVVP in big ET-1), which have previously been demonstrated to be essential for eliciting efficient hydrolysis of the W21−V22 bond in big ET-1 and which have the sequence QTVP in big ET-3. Differences in affinity between big ET-1 and big ET-3 for ECE-1 thus appear to be due solely to sequence variations in the local region of the cleavage site.