Profiling Primary Protease Specificity by Peptide Synthesis on a Solid Support

Abstract

Screening umgekehrt: Das Aufzeichnen der Fluoreszenz während der Protease‐katalysierten Kupplung von Aminosäuren auf einem festen Träger vereinfacht das primäre Screening von Protease‐Spezifität erheblich gegenüber dem Verfolgen einer Peptidhydrolyse (siehe Bild, AA=Aminosäure). Auf diesem Weg sollte eine flexible und schnelle Hochdurchsatz‐Identifizierung und ‐Charakterisierung von Proteasen ohne teuer markierte Peptidarrays möglich werden.

Enhancement of the enzymatic digestibility of sugarcane bagasse by steam pretreatment impregnated with hydrogen peroxide

“…Peptide synthesis catalysed by proteases on solid phase has been largely studied by the group of Flitsch. PEGA 1900 was demonstrated to be fully accessible to chymotrypsin and thermolysin and both peptidases can be used for efficient peptide synthesis or hydrolysis 66…”

“…The group of Flitsch reported a primary screening of protease specificity carried out by reverse peptide hydrolysis (i.e. peptide synthesis) starting from amino acids anchored on PEGA 1900 and by using fluorescence‐labelled amino acids for the coupling step 66. The experiments were developed on a 96‐well microtitre and the specificity of the proteases was indicated by the fluorescence patterns.…”

“…The experiments were developed on a 96‐well microtitre and the specificity of the proteases was indicated by the fluorescence patterns. Moreover, an array of glycopeptides (chemically synthesised on solid phase) were used as substrates to evaluate and compare the ability of the two proteases to hydrolyse the peptide bond in the proximity of N‐glycans 66…”

“…This technology has found a promising route in the use of biocatalyst for the development of highly selective assays under mild operative conditions. For example, methods for screening protease selectivity have been developed 66, 77…”