Abstract
We report for the first time single bead spatially resolved activity measurements of solid-phase biocatalytic systems followed in real-time. Trypsin cleavage of Bz-Arg-OH and subtilisin cleavage of Z-Gly-Gly-Leu-OH each liberate a free amino group on aminocoumarin covalently immobilised to PEGA1900 beads [a co-polymer of poly(ethylene glycol) with molecular mass of 1900 cross-linked with acrylamide]. This restores fluorescence which is imaged in optical sections by two-photon microscopy. For trypsin cleavage, fluorescence is restricted initially to surface regions, with more than 1 hour needed before reaction is fully underway in the bead centre, presumably reflecting slow enzyme diffusion. In contrast, for subtilisin cleavage fluorescence develops throughout the bead more quickly.
Original language | English |
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Pages (from-to) | 1321-1326 |
Number of pages | 6 |
Journal | Advanced Synthesis and Catalysis |
Volume | 349 |
Issue number | 8-9 |
DOIs | |
Publication status | Published - 2007 |
Keywords
- protease cleavage
- two-photon microscopy
- spatial resolution
- solid-phase biocatalysis
- real-time imaging