Bifunctional analogues of SFTI-1 found to be resistant to hydrolysis by trypsin.
The experiments were conducted at 25Â¬C in 50 mM sodium acetate buffer containing 20 mM CaCl2, pH 3.5. Solutions of peptide and trypsin were mixed together so that the final concentration of the peptide was 200 Â¬ÂµM and that of trypsin 2 Â¬ÂµM (1 mole%). At discrete intervals, a 195 Â¬Âµl aliquot was taken and the reaction stopped by addition of 5 Â¬Âµl of TFA. The samples were then analysed directly by RP-HPLC.
When hydrolysis of the peptides was followed at pH 8 using catalytic quantities of trypsin (1 mol%), none of the peptides showed significant levels of hydrolysis products until at least 30 min of incubation, as judged by HPLC analysis from the still intact peak of the original inhibitors. This shows that all of these synthetic peptides are significantly resistant to hydrolysis by trypsin.