Fluorescence-Based On-Resin Detection of Three Model Proteases

A new approach to on-resin detection of three model proteases (trypsin, chymotrypsin, and thrombin) has been developed, while at the same time already described methodology for simultaneous two enzymes (trypsin chymotrypsin) additionally generalized. Appropriate immobilized substrates, comprising specifically cleavable peptide sequences capped with fluorescent dyes, have synthesized on Rink Amide PEGA resin or Amino modified backbone amide linker (BAL). Resulting solid support-bound probes were then dispersed into Tris-HCl buffer solution (pH = 8.0) subjected enzymatic cleavage. Liberated fluorophores tracked by fluorescence measuring. The competitive activities studied towards thrombin probe efficiently limited controlled employing a Bowman-Birk inhibitor system.