Exponential feeding strategy development for benzaldehyde lyase production by recombinant "Escherichia coli"

Taşpınar, Hatice
In this study, the aim was to investigate the effects of exponential feeding strategy on benzaldehyde lyase (BAL) production by recombinant Escherichia coli BL21. For this purpose, the effects of medium components were investigated to optimize the initial medium composition of the fed-batch fermentations. For the batch bioreactor operations, the highest cell concentration and BAL activity were achieved in a media containing 30 g L-1 pretreated molasses, and 5 g L-1 (NH4)2HPO4 as 5.07 g L-1, and 1611 U ml-1 at t=8 h, respectively. Thereafter, in order to increase the cell growth and BAL production while avoiding acetate accumulation, fed-batch bioreactor operations were conducted with exponential feeding at different specific growth rates namely, 0.1 h-1 (mu-0.1), 0.15 h-1 (mu-0.15), and 0.2 h-1 (mu-0.2), and a combined exponential and constant feeding (mu-0.2+) strategy. In the experiments, 9 hours of batch-wise operation with the optimized production medium was followed by a fed-batch operation phase using the pre-determined exponential feeding profiles and for mu-0.2+ operation after 10 hours of exponential feeding as mu-0.2, where the feed rate was kept constant at 21.6 g h-1. Additionally, the plasmid stability was investigated using the feeding method of mu-0.2+ operation with antibiotics in the feed solution, and it was observed that the plasmid was stable. Among the three exponential feeding conditions, the highest cell concentration and BAL activity were determined in -0.15 operation as 21.7 g L−1 and 8379 U ml-1, respectively. In mu-0.2+ operation, the maximum cell concentration was achieved as 28.1 g L-1 which was higher than mu-0.15 experiment, though a lower maximum BAL activity was obtained as 6695 U ml-1.


Influence of oxygen transfer on benzaldehyde lyase production by recombinant Escherichia coli BL21(DE3) pLySs
Angardi, Vahideh; Çalık, Pınar; Department of Chemical Engineering (2007)
In this study, the effects of oxygen transfer conditions on the synthesis of the enzyme benzaldehyde lyase as intracellular in recombinant E. coli BL21 (DE3) pLysS was investigated sistematically and a comprehensive model was developed to determine benzaldehyde lyase activity. For this purpose, the research program was carried out in mainly two parts. In the first part of study, the effects of oxygen transfer together with the mass transfer coefficient (KLa), enhancement factor E (=KLa/KLao), volumetric oxy...
Comparison of benzaldehyde lyase production capacity in recombinant Escherichia coli and recombinant Bacillus species
Kaya, Hande; Çalık, Pınar; Department of Chemical Engineering (2006)
In this study, the benzaldehyde lyase (BAL, EC production in E. coli BL21 (DE3) pLySs as intracellular and in Bacillus species as extracellular were investigated, and comparison of the production capacity of the enzyme in the developed recombinant microorganisms were compared. For this purpose, firstly, PCR amplified bal gene was cloned into pRSETA vector which is under the control of strong T7 promoter and expressed in E. coli BL21 (DE3) pLysS strain. With developed recombinant E. coli BL21 (DE3)...
Molecular cloning and co-expression of Thermoplasma volcanium proteasome subunit genes
Kocabıyık, Semra; Zwickl, Peter; Ozdogan, Seda (Elsevier BV, 2010-10-01)
In this study we describe, the construction of a co-expression vector allowing simultaneous production of Thermoplasma volcanium 20S proteasome alpha- and beta-subunits in Escherichia coli. This heterologous expression system provided high level production of fully active 205 proteasome that can be purified easily by using a conventional two-step chromatographic technique. The recombinant proteasome was purified to homogeneity 12-fold with a specific activity of 26.5 U/mg. Sodium dodecyl sulfate-polyacrylam...
Recombinant therapeutic protease production by Bacillus sp.
Korkmaz, Nuriye; Çalık, Pınar; Department of Chemical Engineering (2007)
The first aim of this study is the development of extracellular recombinant therapeutic protease streptokinase producing Bacillus sp., and the second aim is to determine fermentation characteristics for streptokinase production. In this context, the signal (pre-) DNA sequence of B.licheniformis (DSM1969) extracellular serine alkaline protease enzyme gene (subC: Acc. No. X03341) was ligated to 5’ end of the streptokinase gene (skc: Acc. No. S46536) by SOE (Gene Splicing by Overlap Extension) method through P...
Bioprocess parameters and oxygen transfer characteristics in beta-lactamase production by Bacillus species
Celik, E; Çalık, Pınar (Wiley, 2004-03-01)
After screening potential beta-lactamase producers in a medium containing penicillin G, an inducible (Bacillus subtilis NRS 1125) and a constitutive (Bacillus licheniformis 749/C ATCC 25972) P-lactamase producer were selected. As the highest enzyme activity was obtained with B. licheniformis 749/C, the effects of the concentration of carbon sources, i.e., glucose, fructose, sucrose, citric acid, and glycerol, and nitrogen sources, i.e., (NH4)(2)HPO4, NH4Cl, yeast extract, casamino acids and peptone, pH, and...
Citation Formats
H. Taşpınar, “Exponential feeding strategy development for benzaldehyde lyase production by recombinant “Escherichia coli”,” M.S. - Master of Science, Middle East Technical University, 2010.