High volumetric yields of functional dimeric miniantibodies in Escherichia coli, using an optimized expression vector and high-cell-density fermentation under non-limited growth conditions

Applied Microbiology and Biotechnology
U HornD Riesenberg

Abstract

Functional bivalent miniantibodies, directed against the epidermal growth factor receptor, accumulated to more than 3 gl-1 in high-cell-density cultures of Escherichia coli RV308(pHKK) on a pilot scale. The miniantibodies consist of scFv fragments with a C-terminal hinge followed by a helix-turn-helix motif, which homodimerizes in vivo. The improved expression vector pHKK is characterized by the hoklsok suicide system, improving plasmid maintenance, and the inducible lac pl o promoter system with the very strong T7g10 Shine-Dalgarno sequence. The expression unit is flanked by terminators. The prototrophic RV308 cells were cultivated in glucose mineral salt medium and reached a cell density of 145 g dry biomass l-1 after 33 h. After induction, growth continued almost unchanged for a further 4 h with concomitant miniantibody formation. In the fedbatch phase, the concentration of glucose was kept almost constant at the physiological level of approximately 1.5 gl-1, using on-line flow injection analysis for control. Surprisingly, E. coli RV308(pHKK) did not accumulate significant amounts of the metabolic by-product acetate under these unlimited aerobic growth conditions.

Citations

Jul 14, 2010·Applied Microbiology and Biotechnology·Ján KrahulecJana Machálková
Dec 10, 2009·Journal of Industrial Microbiology & Biotechnology·Thi Thuy TranRajni Hatti-Kaul
May 19, 2005·Biotechnology Advances·Joseph Shiloach, Rephael Fass
Feb 12, 2004·Journal of Immunological Methods·Fabrice Le GallSergey M Kipriyanov
Jun 1, 2005·Protein Expression and Purification·Fatemeh RahbarizadehAbdol-Amir Allameh
Aug 26, 1998·FEBS Letters·K M MüllerA Plückthun
Aug 25, 2001·Journal of Chromatography. a·T K NevanenT T Teeri
Feb 27, 2001·Journal of Immunological Methods·P Carter
Aug 8, 2002·Journal of Immunological Methods·Dan LuZhenping Zhu
Jan 29, 2000·Journal of Immunological Methods·C SchaffitzelA Plückthun
Aug 1, 2000·Immunology Today·M LittleG Moldenhauer
Mar 21, 2001·Vaccine·K PeetersA Depicker
Oct 3, 2002·Protein Expression and Purification·Michael A ThielKeryn A Williams
May 10, 2013·ACS Synthetic Biology·Felix MoserChristopher A Voigt
Oct 4, 2012·Preparative Biochemistry & Biotechnology·Mustafa KahramanCandan Tamerler
May 7, 2004·Applied and Environmental Microbiology·K J MukherjeeD K Summers
Mar 24, 2005·Bioscience, Biotechnology, and Biochemistry·Dae-Hee LeeBum-Shik Hong
Jun 1, 1997·Immunotechnology : an International Journal of Immunological Engineering·A Plückthun, P Pack
Mar 21, 2001·Biotechnology and Bioengineering·M AkessonJ P Axelsson
Feb 20, 1998·Microbiological Research·J L Mörbe, D Riesenberg
Nov 4, 2000·Microbiological Research·V SchroeckhD Riesenberg
Jan 24, 2009·Journal of Biotechnology·Thomas I PotgieterMarc d'Anjou
Sep 4, 2007·Journal of Molecular Biology·Georg ZocherGeorg E Schulz

❮ Previous
Next ❯

Related Concepts

Trending Feeds

COVID-19

Coronaviruses encompass a large family of viruses that cause the common cold as well as more serious diseases, such as the ongoing outbreak of coronavirus disease 2019 (COVID-19; formally known as 2019-nCoV). Coronaviruses can spread from animals to humans; symptoms include fever, cough, shortness of breath, and breathing difficulties; in more severe cases, infection can lead to death. This feed covers recent research on COVID-19.

Blastomycosis

Blastomycosis fungal infections spread through inhaling Blastomyces dermatitidis spores. Discover the latest research on blastomycosis fungal infections here.

Nuclear Pore Complex in ALS/FTD

Alterations in nucleocytoplasmic transport, controlled by the nuclear pore complex, may be involved in the pathomechanism underlying multiple neurodegenerative diseases including Amyotrophic Lateral Sclerosis and Frontotemporal Dementia. Here is the latest research on the nuclear pore complex in ALS and FTD.

Applications of Molecular Barcoding

The concept of molecular barcoding is that each original DNA or RNA molecule is attached to a unique sequence barcode. Sequence reads having different barcodes represent different original molecules, while sequence reads having the same barcode are results of PCR duplication from one original molecule. Discover the latest research on molecular barcoding here.

Chronic Fatigue Syndrome

Chronic fatigue syndrome is a disease characterized by unexplained disabling fatigue; the pathology of which is incompletely understood. Discover the latest research on chronic fatigue syndrome here.

Evolution of Pluripotency

Pluripotency refers to the ability of a cell to develop into three primary germ cell layers of the embryo. This feed focuses on the mechanisms that underlie the evolution of pluripotency. Here is the latest research.

Position Effect Variegation

Position Effect Variagation occurs when a gene is inactivated due to its positioning near heterochromatic regions within a chromosome. Discover the latest research on Position Effect Variagation here.

STING Receptor Agonists

Stimulator of IFN genes (STING) are a group of transmembrane proteins that are involved in the induction of type I interferon that is important in the innate immune response. The stimulation of STING has been an active area of research in the treatment of cancer and infectious diseases. Here is the latest research on STING receptor agonists.

Microbicide

Microbicides are products that can be applied to vaginal or rectal mucosal surfaces with the goal of preventing, or at least significantly reducing, the transmission of sexually transmitted infections. Here is the latest research on microbicides.