D-AMINO-ACID SCAN OF ENDOTHELIN - IMPORTANCE OF AMINO-ACIDS ADJACENT TO CYSTEINYL RESIDUES IN ISOMERIC SELECTIVITY

A systematic approach to map the functionally important determinants o f endothelin-1 (ET-1) by a D-amino acid scan is described. Correct ori entation of the amino acid side chains was generally of paramount impo rtance both for binding at the ET(A) receptor and for contracting acti vity This was particularly valid for positions 2, 8, 14, 16-21 (the fo ur Cys residues were kept unaltered). Nevertheless, increment of bindi ng affinity was observed by inversion of configuration at positions 6, 7, 9 and 10. In addition, [D-Lys(9)]ET was an agonist about four time s more potent than the natural compound. Usually both 1,4- and 1,3-iso mers (corresponding, respectively to the correct and misfolded disulfi de bridges of ET) were obtained, and usually the isomer formed in larg er amount had the higher HPLC retention time and the higher biological activity. However, four out of seventeen single-point D-amino acid an alogues could he isolated only in one isomeric form. In three cases (D -Ser(2), D-Ser(4), D-Val(12)), the inverted amino acid was adjacent to a Cys residue, and in one case (D-Lys(9)) it was one amino acid apart , thus suggesting a possible effect of the bridged cysteinyl residues in isomeric selectivity.