A sensitive fluorometric assay for tissue transglutaminase

We have devised a highly sensitive fluorometric well plate assay for tissue transglutaminase that is suitable for multiple kinetic analyses/high-throu ghput screening of chemical inventories for inhibitors of this enzyme. The procedure measures the rate of fluorescence enhancement (lambda (exc) 260 n m, lambda (em) 538 nm) when 1-N-(carbobenzoxy-L-glutaminylglycyl)-5-N-(5'N' N'-dimethylaminonaphthalenesulfonyl) diamidopentane (glutaminyl substrate) is cross-linked to dansyl cadaverine (amine substrate). The assay procedure can be used to measure the activity of as little as 60 muU Of purified gui nea pig liver tissue transglutaminase (4.2 ng or 54 fmol of enzyme). (C) 20 01 Academic Press.