Receptor binding activity and cytosolic free calcium response by synthetic endothelin analogs in cultured rat vascular smooth muscle cells

Using a variety of synthetic analogs of porcine endothelin (pET), we have studied the effects of these analogs on receptor binding activity and cytosolic free Ca 2+ concentrations ([Ca 2+] i) in cultured rat vascular smooth muscle cells (VSMC). Removal of C-terminal Trp 21 residue, truncated derivatives pET(1–15) and (16–21), substitution of disulfide bond, Cys(3–11) or Cys(1–15), by Cys (Acm), all resulted in a complete loss of receptor binding activity and [Ca 2+] i response, while N-terminal elongation of Lys-Arg residues, but not oxidation of Met 7 residue decreased receptor binding activity and [Ca 2+] i response. [Cys 1–15, Cys 3–11]pET was far more potent than [Cys 1–11, Cys 3–15]pET in receptor binding and [Ca 2+] i response. These data indicate that the C-terminal Trp 21 as well as the proper double cyclic structure formed by the intramolecular disulfide bonds of the pET molecule are essential for receptor binding and subsequent [Ca 2+] i increase in rat VSMC.