A new proteinase 3 substrate with improved selectivity over human neutrophil elastase.
We report the synthesis and enzymatic studies on a new proteinase 3 intermolecular quenched substrate with enhanced selectivity over neutrophil elastase. Using combinatorial chemistry methods, we were able to synthesize the hexapeptide library with the general formula ABZ-Tyr-Tyr-Abu-X1'-X2'-X3'-Tyr(3-NO2)-NH2 using the mix and split method. The iterative deconvolution of such a library allowed us to obtain the sequence ABZ-Tyr-Tyr-Abu-Asn-Glu-Pro-Tyr(3-NO2)-NH2 with a high specificity constant (kcat/KM=1534×10(3)M(-1)s(-1)) and superior selectivity over neutrophil elastase and other neutrophil-derived serine proteases. Moreover, using the obtained substrate, we were able to detect a picomolar concentration of proteinase 3 (PR3). Incubation of the above-mentioned substrate with neutrophil lysate resulted in a strong fluorescent signal that was significantly reduced in the presence of a PR3 selective inhibitor.
['Popow-Stellmaszyk|J|J|', 'Wysocka|M|M|', 'Lesner|A|A|', 'Korkmaz|B|B|', 'Rolka|K|K|']
D020650:Combinatorial Chemistry Techniques D006801:Humans D019272:Leukocyte Elastase D053146:Myeloblastin D009842:Oligopeptides D013379:Substrate Specificity
Proteinase 3 Intermolecular quenched substrate Combinatorial chemistry Neutrophil serine proteases