Scorpion antivenom nanobody production in Escherichia coli (Recombinant bispecific VHH)

Scorpionism is a serious public health problem in tropical areas, especially in Africa, southern India, the Middle East, and Latin America. There are about 12 species of scorpions that are dangerous to humans and all belong to the Buthidae family. In particular, venom on scorpion Androctonus australis hector is particularly toxic to humans. Serotherapy uses antibodies or fragments of antibodies to target the neurotoxins of the scorpion venom. Nanobodies, camelid-derived antibodies, are more ...

Programme: This Project is not associated with a Programme

Public web page: Not specified

Start date: 6th Feb 2017

Characterization of E. coli WK6 strain under aerobic conditions
No description specified

Submitter: Susana Alonso Villela

Investigation: Effect of the induction temperature in nanobody...

Assays: Fed-batch culture of WK6 strain, without induction - BR01

Fed-batch culture of WK6 strain, induction at 29°C - BR09

5L bioreactor culture of reference strain E. coli WK6. Protein induction promoter when biomass reached 25 g cdw/L and at 29°C for 28 h

Submitter: Susana Alonso Villela

Assay type: Cultivation Experiment

Technology type: Fed-batch

Investigation: Effect of the induction temperature in nanobody...

Study: Characterization of E. coli WK6 strain under in...

Fed-batch culture of NbF12-10 strain, induction at 29°C - BR08

5L bioreactor culture of recombinant strain E. coli NbF12-10. Protein induction promoter when biomass reached 30 g cdw/L and at 29°C for 37 h

Submitter: Susana Alonso Villela

Assay type: Cultivation Experiment

Technology type: Fed-batch

Investigation: Effect of the induction temperature in nanobody...

Study: Recombinant nanobody CH10-12 and NbF12-10 produ...

Fed-batch culture of CH10-12 strain, induction at 32°C - BR07

5L bioreactor culture of recombinant strain E. coli CH10-12. Protein induction promoter when biomass reached 30 g cdw/L and at 32°C for 35 h

Submitter: Susana Alonso Villela

Assay type: Cultivation Experiment

Technology type: Fed-batch

Investigation: Effect of the induction temperature in nanobody...

Study: Recombinant nanobody CH10-12 and NbF12-10 produ...

Fed-batch culture of CH10-12 strain, induction at 29°C - BR06

5L bioreactor culture of recombinant strain E. coli CH10-12. Protein induction promoter when biomass reached 30 g cdw/L and at 29°C for 38 h

Submitter: Susana Alonso Villela

Assay type: Cultivation Experiment

Technology type: Fed-batch

Investigation: Effect of the induction temperature in nanobody...

Study: Recombinant nanobody CH10-12 and NbF12-10 produ...

Fed-batch culture of CH10-12 strain, induction at 37°C - BR05

5L bioreactor culture of recombinant strain E. coli CH10-12. Protein induction promoter when biomass reached 25 g cdw/L and at 37°C for 6 h

Submitter: Susana Alonso Villela

Assay type: Cultivation Experiment

Technology type: Fed-batch

Investigation: Effect of the induction temperature in nanobody...

Study: Recombinant nanobody CH10-12 production at diff...

Fed-batch culture of CH10-12 strain, induction at 33°C - BR04

5L bioreactor culture of recombinant strain E. coli CH10-12. Protein induction promoter when biomass reached 25 g cdw/L and at 33°C for 6 h

Submitter: Susana Alonso Villela

Assay type: Cultivation Experiment

Technology type: Fed-batch

Investigation: Effect of the induction temperature in nanobody...

Study: Recombinant nanobody CH10-12 production at diff...

Fed-batch culture of CH10-12 strain, induction at 28°C - BR02

5L bioreactor culture of recombinant strain E. coli CH10-12 in a batch/fed-batch strategy at 37°C (BR02). Protein induction promoted when biomass reached 25 g cdw/L and at 28°C for 12 h

Submitter: Susana Alonso Villela

Assay type: Cultivation Experiment

Technology type: Fed-batch

Investigation: Effect of the induction temperature in nanobody...

Study: Recombinant nanobody CH10-12 production at diff...

Fed-batch culture of WK6 strain, without induction - BR01

5L bioreactor culture of strain E. coli WK6 in a batch/fed-batch strategy at 37°C. Characterization of biomass growth and oxygen transfer

Submitter: Susana Alonso Villela

Assay type: Cultivation Experiment

Technology type: Fed-batch

Investigation: Effect of the induction temperature in nanobody...

Study: Characterization of E. coli WK6 strain under ae...

BR01 samples

Samples taken in the 5L bioreactor BR01 in the culture of E. coli WK6 in batch and fed-batch strategy

Creator: Susana Alonso Villela

Submitter: Susana Alonso Villela

Cultivation of P. putida (and E. coli) in rich-, minimal-, and selective medium
No description specified

Creator: Rita Volkers

Submitter: Rita Volkers

Mix&Go competent cells and transformation of E. coli
No description specified

Creator: Rita Volkers

Submitter: Rita Volkers

Colony PCR of S. cerevisiae, P. putida, E. coli
No description specified

Creator: Rita Volkers

Submitter: Rita Volkers

Making electro-competent E. coli cells and transformation of them
No description specified

Creator: Rita Volkers

Submitter: Rita Volkers

Plasmid isolation from E. coli and P. putida using the Thermo Scientific GeneJET Plasmid Miniprep Kit
No description specified

Creator: Rita Volkers

Submitter: Rita Volkers

Making a growth curve of a bacterial or yeast strain
No description specified

Creator: Rita Volkers

Submitter: Rita Volkers

Effect of temperature on the production of a recombinant antivenom in fed-batch mode

Abstract (Expand)

In the pharmaceutical industry, nanobodies show promising properties for its application in serotherapy targeting the highly diffusible scorpion toxins. The production of recombinant nanobodies in … Escherichia coli has been widely studied in shake flask cultures in rich medium. However, there are no upstream bioprocess studies of nanobody production in defined minimal medium and the effect of the induction temperature on the production kinetics. In this work, the effect of the temperature during the expression of the chimeric bispecific nanobody CH10-12 form, showing high scorpion antivenom potential, was studied in bioreactor cultures of E. coli. High biomass concentrations (25 g cdw/L) were achieved in fed-batch mode, and the expression of the CH10-12 nanobody was induced at temperatures 28, 29, 30, 33, and 37°C with a constant glucose feed. For the bispecific form NbF12-10, the induction was performed at 29°C. Biomass and carbon dioxide yields were reported for each culture phase, and the maintenance coefficient was obtained for each strain. Nanobody production in the CH10-12 strain was higher at low temperatures (lower than 30°C) and declined with the increase of the temperature. At 29°C, the CH10-12, NbF12-10, and WK6 strains were compared. Strains CH10-12 and NbF12-10 had a productivity of 0.052 and 0.021 mg/L/h of nanobody, respectively, after 13 h of induction. The specific productivity of the nanobodies was modeled as a function of the induction temperature and the specific growth rates. Experimental results confirm that low temperatures increase the productivity of the nanobody. Key points • Nanobodies with scorpion antivenom activity produced using two recombinant strains. • Nanobodies production was achieved in fed-batch cultures at different induction temperatures. • Low induction temperatures result in high volumetric productivities of the nanobody CH10-12.

Authors: Susana María Alonso Villela, Hazar Ghezal-Kraïem, Balkiss Bouhaouala-Zahar, Carine Bideaux, César Arturo Aceves Lara, Luc Fillaudeau

Date Published: 1st Feb 2021

Publication Type: Journal

MFCS BioPAT - BR04

5L bioreactor of E. coli CH10-12 in a batch/fed-batch strategy at 37°C, induction promoted for 6h at 33°C MFCS BioPAT Sartorius

MFCS BioPAT - BR03

5L bioreactor of E. coli CH10-12 in a batch/fed-batch strategy at 37°C, induction promoted for 10h at 30°C MFCS BioPAT Sartorius

MFCS BioPAT - BR02

5L bioreactor of E. coli CH10-12 in a batch/fed-batch strategy at 37°C, induction promoted for 12h at 28°C MFCS BioPAT Sartorius

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