Abstract
Heat conditioning of cell homogenates of B. cereus and a recombinant E. coli was studied for the isolation of leucine dehydrogenase and alanine racemase, respectively. The strain of E. coli carried the gene of the thermostable alanine racemase from B. stearothermophilus. Activity loss can be minimized (<5%) and aggregation and flocculation of soluble proteins (70–80%) and other cell components can be achieved, depending on temperature, biomass concentration and pH-value.
Thereby a 3–6 fold increase in specific activity was obtained. The resulting extract after solid-liquid separation showed lower viscosity and less turbidity than unheated controls, making it more suitable for chromatographic separations.
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Abbreviations
- ADH :
-
alcohol dehydrogenase
- AlaR :
-
alanine racemase
- BM %:
-
biomass concentration
- D 1/s:
-
Shear rate
- η mPas:
-
viscosity
- EF :
-
enrichment factor
- LeuDH :
-
leucine dehydrogenase
- \(\dot Q\) KW:
-
heat rate
- RA %:
-
remaining activity
- RCF :
-
relative centrifugal force
- RP %:
-
remaining protein
- ϱ kg/dm3 :
-
density of homogenate
- T F°C:
-
temperature of feed
- T HI°C:
-
temperature of heating fluid (inlet)
- T HO°C:
-
temperature of heating fluid (outlet)
- K Pdm3/h:
-
volumetric flow rate of the feed
References
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Kula, MR., Schnell, J. Investigations of heat treatment to improve the isolation of intracellular enzymes. Bioprocess Eng. 5, 31–38 (1990). https://doi.org/10.1007/BF00369644
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DOI: https://doi.org/10.1007/BF00369644