Robert Huber and Helene Wulfhorst contributed equally to this work.
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Process Sensing and Control
Screening for enzyme activity in turbid suspensions with scattered light
Article first published online: 7 FEB 2011
DOI: 10.1002/btpr.519
Copyright © 2011 American Institute of Chemical Engineers (AIChE)
Additional Information
How to Cite
Huber, R., Wulfhorst, H., Maksym, L., Stehr, R., Pöhnlein, M., Jäger, G., Spieß, A. C. and Büchs, J. (2011), Screening for enzyme activity in turbid suspensions with scattered light. Biotechnol Progress, 27: 555–561. doi: 10.1002/btpr.519
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Robert Huber and Helene Wulfhorst contributed equally to this work.
Publication History
- Issue published online: 11 APR 2011
- Article first published online: 7 FEB 2011
- Accepted manuscript online: 11 OCT 2010 08:40AM EST
- Manuscript Revised: 14 JUL 2010
- Manuscript Received: 11 MAR 2010
Funded by
- Stiftung Industrieforschung, Köln, Germany. Grant Number: Project S735
- Excellence Initiative by the German federal and state governments to promote science and research at German universities
- Abstract
- Article
- References
- Cited By
Keywords:
- high throughput;
- protein hydrolysis;
- microtiter plate;
- turbid suspension;
- protease activity
Abstract
New screening techniques for improved enzyme variants in turbid media are urgently required in many industries such as the detergent and food industry. Here, a new method is presented to measure enzyme activity in different types of substrate suspensions. This method allows a semiquantitative determination of protease activity using native protein substrates. Unlike conventional techniques for measurement of enzyme activity, the BioLector technology enables online monitoring of scattered light intensity and fluorescence signals during the continuous shaking of samples in microtiter plates. The BioLector technique is hereby used to monitor the hydrolysis of an insoluble protein substrate by measuring the decrease of scattered light. The kinetic parameters for the enzyme reaction (Vmax,app and Km,app) are determined from the scattered light curves. Moreover, the influence of pH on the protease activity is investigated. The optimal pH value for protease activity was determined to be between pH 8 to 11 and the activities of five subtilisin serine proteases with variations in the amino acid sequence were compared. The presented method enables proteases from genetically modified strains to be easily characterized and compared. Moreover, this method can be applied to other enzyme systems that catalyze various reactions such as cellulose decomposition. © 2011 American Institute of Chemical Engineers Biotechnol. Prog., 2011