Relation between cell disruption conditions, cell debris particle size, and inclusion body release

Van Hee, P., Middelberg, A. P. J., Van der Lans, R. G. J. M. and Van der Wielen, L. A. M. (2004) Relation between cell disruption conditions, cell debris particle size, and inclusion body release. Biotechnology and Bioengineering, 88 1: 100-110. doi:10.1002/bit.20215


Author Van Hee, P.
Middelberg, A. P. J.
Van der Lans, R. G. J. M.
Van der Wielen, L. A. M.
Title Relation between cell disruption conditions, cell debris particle size, and inclusion body release
Journal name Biotechnology and Bioengineering   Check publisher's open access policy
ISSN 0006-3592
Publication date 2004
Sub-type Article (original research)
DOI 10.1002/bit.20215
Volume 88
Issue 1
Start page 100
End page 110
Total pages 11
Editor D. S. Clark
Place of publication Hoboken
Publisher John Wiley & Sons
Collection year 2004
Language eng
Subject C1
290699 Chemical Engineering not elsewhere classified
670499 Other
Abstract The efficiency of physical separation of inclusion bodies from cell debris is related to cell debris size and inclusion body release and both factors should be taken into account when designing a process. In this work, cell disruption by enzymatic treatment with lysozyme and cellulase, by homogenization, and by homogenization with ammonia pretreatment is discussed. These disruption methods are compared on the basis of inclusion body release, operating costs, and cell debris particle size. The latter was measured with cumulative sedimentation analysis in combination with membrane-associated protein quantification by SDS-PAGE and a spectrophotometric pepticloglycan quantification method. Comparison of the results obtained with these two cell debris quantification methods shows that enzymatic treatment yields cell debris particles with varying chemical composition, while this is not the case with the other disruption methods that were investigated. Furthermore, the experiments show that ammonia pretreatment with homogenization increases inclusion body release compared to homogenization without pretreatment and that this pretreatment may be used to control the cell debris size to some extent. The enzymatic disruption process gives a higher product release than homogenization with or without ammonia pretreatment at lower operating costs, but it also yields a much smaller cell debris size than the other disruption process. This is unfavorable for centrifugal inclusion body purification in this case, where cell debris is the component going to the sediment and the inclusion body is the floating component. Nevertheless, calculations show that centrifugal separation of inclusion bodies from the enzymatically treated cells gives a high inclusion body yield and purity. (C) 2004 Wiley Periodicals, Inc.
Keyword Biotechnology & Applied Microbiology
Cell Debris Particle Size
Pseudomonas Putida
Chemical Lysis
Homogenization
Polyhydroxyalkanoate
Inclusion Body
High-pressure Homogenizer
Escherichia-coli
Bakers-yeast
Muramic Acid
Lactic-acid
Proteins
Disintegration
Peptidoglycan
Separation
Extraction
Q-Index Code C1

 
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Created: Wed, 15 Aug 2007, 03:12:40 EST