Semi-defined medium based feeding strategy development for human growth hormone production by recombinant bacillus subtilis/

Download
2014
Öztürk, Sibel
In this study, the aim was to develop an efficient semi-defined medium based continuous feeding strategy for recombinant human growth hormone (rhGH) production by Bacillus subtilis 1A178 scoC knockout (scoC-) strain carrying pMK4::pre(subC)::hGH plasmid. In this context, firstly laboratory-scale air-filtered shake bioreactor experiments were carried out in order to investigate the effects of peptone as the complex carbon and nitrogen source on the cell growth and rhGH production both in the absence and the presence of MgSO4. The highest cell and rhGH concentrations were achieved as CX=1.92 g L-1 and CrhGH=140 mg L-1, respectively, at t=24 h in the semi-defined medium PM-8, containing 8 g L-1 peptone and 0.122 g L-1 MgSO4 in addition to the defined production medium consisting of 8 g L-1 glucose, 4.71 g L-1 (NH4)2HPO4, 2 g L-1 KH2PO4, 0.043 g L-1 Na2HPO4, 5.63 g L-1 NaH2PO4, and 5 ml L-1 trace salt solution (PTM1). Based on these results, in the second phase of the research, MgSO4-containing production medium was used for pilot-scale bioreactor experiments together with the feed substrate stock solutions containing peptone at two different concentrations. Six continuous feeding strategies were designed for semi-batch rhGH production process based on the pre-determined specific growth rates, μ0=0.10, 0.17 and 0.25 h-1, and the transition cultivation times from batch to semi-batch operation, tT=2 h and tT=4 h, were investigated. The highest rhGH concentration was obtained as CrhGH=497 mg L-1 in the semi-defined medium based feeding strategy conducted at μ0=0.25 h-1 where the transition cultivation time was tT=4 h by using the feed substrate stock solution comprising of 200 g L-1 glucose, 117.65 g L-1 (NH4)2HPO4, 100 g L-1 peptone and 5 mL L-1 PTM1 at t=22 h when the cell concentration was reached to CX=8.29 g L-1. The overall product and cell yield on total substrate were attained as Y'(P/S)=7.21 g kg-1 and Y'(X/S)=0.12 g g-1, respectively.

Suggestions

Human growth hormone production by metabolically engineered b. subtilis : feeding strategy development
Gökbudak, Burcu; Çalık, Pınar; Özdamar, Tunçer H.; Department of Chemical Engineering (2013)
In this study, the aim is to develop an effective feeding strategy for therapeutically important protein, recombinant human growth hormone (rhGH) production by Bacillus subtilis scoC knockout (scoC-) strain carrying pMK4::pre(subC)::hGH plasmid. In the first part of the research program, laboratory scale air filtered shake bioreactor experiments were conducted to optimize the production conditions. Thereafter, semi-batch production processes at pilot scale bioreactor system were performed by designing expon...
Feeding strategy design for recombinant human growth hormone production by Bacillus subtilis
Sahin, Burcu; Ozturk, Sibel; Çalık, Pınar; Ozdamar, Tuncer H. (2015-10-01)
Defined and semi-defined medium-based feeding strategies were developed to enhance recombinant human growth hormone (rhGH) production by Bacillus subtilis BGSC-1A178 (scoC (-)) strain carrying pMK4::pre(subC)::hGH. Defined medium-based feeding strategies were designed by exponential feeding of glucose and (NH4)(2)HPO4 at two pre-determined specific growth rates, A mu (0) = 0.10 and 0.17 h(-1). Semi-defined medium-based feeding strategies were designed by exponential feeding of substrate solution consisting ...
Defined and complex medium based feeding strategy development for recombinant human growth hormone production by P. pastoris under GAP promoter
Hoxha, Bebeta; Çalık, Pınar; Özdamar, Tunçer H.; Department of Chemical Engineering (2016)
The objective of this study is to investigate different feeding strategies leading to higher recombinant human growth hormone (rhGH) production by Pichia pastoris, driven under constitutive promoter of the glyceraldehyde-3-phosphate dehydrogenase gene. rhGH production took place in a pilot bioreactor where glucose and molasses were examined as carbon sources. For batch phase, which is the first production phase of the reactor, all experiments share the same characteristics where glycerol was utilized as the...
Comparative analysis of product and by-product distributions in defined and complex media in serine alkaline protease production by recombinant Basillus subtilis
Oktar, Ceren; Çalık, Pınar; Department of Chemical Engineering (2003)
In this study, firstly the effects of aspartic acid group amino acids -which were reported to be the potential bottleneck in serine alkaline protease (SAP) synthesis- on SAP production were investigated by substituting at a concentration range of 0-15 mM by using recombinant Bacillus subtilis carrying pHV1434::subC gene. All aspartic acid group amino acids except threonine inhibited SAP activity when CAA= 2.5 mM. The highest SAP activities with asparagine, aspartic acid, lysine, threonine, isoleucine and me...
Exponential feeding strategy development for benzaldehyde lyase production by recombinant "Escherichia coli"
Taşpınar, Hatice; Çalık, Pınar; Özdamar, Tunçer H.; Department of Biotechnology (2010)
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 ...
Citation Formats
S. Öztürk, “Semi-defined medium based feeding strategy development for human growth hormone production by recombinant bacillus subtilis/,” M.S. - Master of Science, Middle East Technical University, 2014.