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Screening of cellulases for biofuel production: Online monitoring of the enzymatic hydrolysis of insoluble cellulose using high‐throughput scattered light detection
Author(s) -
Jäger Gernot,
Wulfhorst Helene,
Zeithammel Erik U.,
Elinidou Efthimia,
Spiess Antje C.,
Büchs Jochen
Publication year - 2011
Publication title -
biotechnology journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.144
H-Index - 84
eISSN - 1860-7314
pISSN - 1860-6768
DOI - 10.1002/biot.201000387
Subject(s) - cellulase , cellulose , cellulosic ethanol , hydrolysis , trichoderma reesei , chemistry , enzymatic hydrolysis , crystallinity , chromatography , chemical engineering , biochemistry , crystallography , engineering
A new prospective cellulase assay simultaneously combining high‐throughput, online analysis and insoluble cellulosic substrates is described. The hydrolysis of three different insoluble cellulosic substrates, catalysed by a commercial cellulase preparation from Trichoderma reesei (Celluclast), was monitored using the BioLector – allowing online monitoring of scattered light intensities in a continuously shaken microtiter plate. Cellulase activities could be quantitatively assayed using the BioLector. At low cellulase/cellulose ratios, the Michaelis‐Menten parameters of the cellulase mixture were mainly affected by the crystallinity index of the cellulose. Here, the apparent maximum cellulase activities inversely correlated with the crystallinity index of the cellulose. At high cellulase/cellulose ratios the particle size of the cellulose, defining the external surface area accessible to the cellulases, was the key determining factor for cellulase activity. The developed technique was also successfully applied to evaluate the pH optimum of cellulases. Moreover, the non‐hydrolytic deagglomeration of cellulose particles was investigated, for the first time, using high‐throughput scattered light detection. In conclusion, this cellulase assay ideally links high‐throughput, online analysis and realistic insoluble cellulosic substrates in one simple system. It will considerably simplify and accelerate fundamental research on cellulase screening.