An echistatin analogue, designated as des(46-49)-[Ala8,37]-echistatin γ, was synthesized chemically by solid-phase peptide synthesis. The analogue was made by replacing Cys8 and Cys37 residues with two alanines and the deletion of C-terminal peptide 46-49 of echistatin γ, resulting in an artificial polypeptide of 45 amino acids with three disulfide bonds. In the platelet aggregation assay, the analogue exhibits almost the same activity as echistatin γ, indicating that the linear sequence of des(46-49)-[Ala8,37]-echistatin γ contains all of the primary-structure information that is required for proper folding of this synthetic polypeptide. The tertiary structure of the analogue, as determined from high-resolution nuclear magnetic resonance (NMR) coupled with dynamic simulated annealing, is very similar to that of echistatin α1 which differs from echistatin γ by 8 residues. In particular the two important sites of the Arg-Gly-Asp (RGD) loop and the C-terminal Lys45, both of which show some degree of disorder, are maintained in similar spatial orientation and proximity as those in echistatin al even without the constraint provided by the disulfide bond of the (Cys8-Cys37) pair. These results provide new insights in further defining distinct structural features of echistatin γ, which are involved in supporting the active polypeptide conformation to achieve biological activity in the absence of one pair of disulfide bonds.
|Number of pages||9|
|Journal||Biochemical and Biophysical Research Communications|
|Publication status||Published - 1996 Mar 18|
ASJC Scopus subject areas
- Molecular Biology
- Cell Biology