Regulatory gene effects on recombinant human growth hormone production by Bacillus subtilis

Şahin, Merve
In this study, regulatory gene effects on recombinant human growth hormone (rhGH) production by Bacillus subtilis were investigated. For this purpose, firstly Bacillus strains, which are deficient in abrB, aprE, degQ, degS, degU, scoC, sinI, sinR, and spo0A genes, were selected according to the regulatory gene network of aprE gene (serine alkaline protease gene of B. subtilis) since due to the degQ promoter and the pre-signal sequence of subC gene cloned in front of the hGH gene, hGH is produced by mimicking the serine alkaline protease synthesis. R-Bacillus strains were constructed by transformation of pMK4::pre(subC)::hGH plasmid to the selected strains. Thereafter, by the laboratory scale experiments, strains having the highest hGH production capacity were determined as scoC, aprE, sinR, and degU knockout strains. Using these strains, fermentation experiments were carried out in pilot-scale bioreactor in defined medium. Effect of pH control was also investigated and the highest cell and hGH concentration was obtained by scoC knockout strain in pH controlled operation as 1.62 kg m-3 and 126 g m-3, respectively. By this strain, the overall product and cell yield on total substrate were found as 16.12 g kg-1 and 0.15 g g-1, respectively. Furthermore, the highest total protease activity was attained by degU knockout strain as 65 U cm-3. On the other hand, maximum total organic acid secretion was determined as 1.31 kg m-3 in aprE knockout strain.


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Arifoğlu, Müge; Çalık, Pınar; Department of Chemical Engineering (2004)
In this study, the effects of oxygen transfer (OT) on beta-lactamase production and on intracellular reaction rates were investigated with Bacillus licheniformis ATCC 2597. In order to clarify the oxygen transfer effects on the production of beta-lactamase, firstly a glucose based defined medium was designed and using this medium, the effects of bioreactor operation parameters, i.e., pH and temperature, on beta-lactamase activity and cell formation were investigated in laboratory scale batch-bioreactors usi...
Codon optimization of xylA gene for recombinant glucose isomerase production in Pichia pastoris and fed-batch feeding strategies to fine-tune bioreactor performance
Ata, Ozge; Boy, Erdem; Gunes, Hande; Çalık, Pınar (Springer Science and Business Media LLC, 2015-05-01)
The objectives of this work are the optimization of the codons of xylA gene from Thermus thermophilus to enhance the production of recombinant glucose isomerase (rGI) in P. pastoris and to investigate the effects of feeding strategies on rGI production. Codons of xylA gene from T. thermophilus were optimized, ca. 30 % of the codons were replaced with those with higher frequencies according to the codon usage bias of P. pastoris, codon optimization resulted in a 2.4-fold higher rGI activity. To fine-tune bio...
Homologous expression of aspartokinase (ask) gene in Streptomyces clavuligerus and its hom-deleted mutant: Effects on cephamycin C production
Özcengiz, Gülay; Ünsaldi, Eser; Taşkin, Bilgin; Liras, Paloma; Piret, Jacqueline (Informa UK Limited, 2010-05-01)
Abstract In this study, the effect of homologous multiple copies of the ask gene, which encodes aspartokinase catalyzing the first step of the aspartate pathway, on cephamycin C biosynthesis in S. clavuligerus NRRL 3585 and its hom mutant was investigated. The intracellular pool levels of aspartate pathway amino acids accorded well with the Ask activity levels in TB3585 and AK39. When compared with the control strain carrying vector alone without any gene insert, amplification of the ask gene in the w...
Fed-Batch Biomolecule Production by Bacillus subtilis: A State of the Art Review
Ozturk, Sibel; Çalık, Pınar; Ozdamar, Tuncer H. (Elsevier BV, 2016-04-01)
Bacillus subtilis is a highly promising production system for various biomolecules. This review begins with the algorithm of fed-batch operations (FBOs) and then illustrates the approaches to design the initial production medium and/or feed stream. Additionally, the feeding strategies developed with or without feedback control for fed-batch B. subtilis fermentations were compiled with a special emphasis on recombinant protein (r-protein) production. For biomolecule production by wild-type B. subtilis, due t...
Differential expression of genes possibly involved in incompatible interaction between barley and powdery mildew
ERSOY, FİGEN; Ridout, Christopher J.; Akkaya, Mahinur (2011-09-01)
The various domains of a plant disease resistance protein from wheat were found to be interacting with yeast proteins when screened via yeast two hybrid analyses. These genes are considered to play roles in disease resistance response. Thus, the expression levels in Mla3 mediated Powdery Mildew (Blumeria graminis f.sp. hordei, Bgh) disease resistance in barley were determined. The barley homologs of ARD1, CPR7, CSE1, GCN2 and SRP72, were partially cloned and sequenced. Their differential expression was conf...
Citation Formats
M. Şahin, “Regulatory gene effects on recombinant human growth hormone production by Bacillus subtilis,” M.S. - Master of Science, Middle East Technical University, 2010.