Cell bank characterization and fermentation optimization for production of recombinant heavy chain C-terminal fragment of botulinum neurotoxin serotype E (rBoNTE(H(c)): antigen E) by Pichia pastoris.

Cell bank characterization and fermentation optimization for production of recombinant heavy chain C-terminal fragment of botulinum neurotoxin serotype E (rBoNTE(H(c)): antigen E) by Pichia pastoris. Research Abstract Details 

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Cell bank characterization and fermentation optimization for production of recombinant heavy chain C-terminal fragment of botulinum neurotoxin serotype E (rBoNTE(H(c)): antigen E) by Pichia pastoris. Abstract Text:

Jayanta Sinha,Mehmet Inan,Sarah Fanders,Shinichi Taoka,Mark Gouthro,Todd Swanson,Rick Barent,Ardis Barthuli,Bonnie M Loveless,Leonard A Smith,Theresa Smith,Ian Henderson,John Ross,Michael M Meagher,

A process was developed for production of a candidate vaccine antigen, recombinant C-terminal heavy chain fragment of the botulinum neurotoxin serotype E, rBoNTE(H(c)) in Pichia pastoris. P. pastoris strain GS115 was transformed with the rBoNTE(H(c)) gene inserted into pHILD4 Escherichia coli-P. pastoris shuttle plasmid. The clone was characterized for genetic stability, copy number, and BoNTE(H(c)) sequence. Expression of rBoNTE(H(c)) from the Mut(+) HIS4 clone was confirmed in the shake-flask, prior to developing a fed-batch fermentation process at 5 and 19 L scale. The fermentation process consists of a glycerol growth phase in batch and fed-batch mode using a defined medium followed by a glycerol/methanol transition phase for adaptation to growth on methanol and a methanol induction phase resulting in the production of rBoNTE(H(c)). Specific growth rate, ratio of growth to induction phase, and time of induction were critical for optimal rBoNTE(H(c)) production and minimal proteolytic degradation. A computer-controlled exponential growth model was used for process automation and off-gas analysis was used for process monitoring. The optimized process had an induction time of 9 h on methanol and produced up to 3 mg of rBoNTE(H(c)) per gram wet cell mass as determined by HPLC and Western blot analysis.

Cell bank characterization and fermentation optimization for production of recombinant heavy chain C-terminal fragment of botulinum neurotoxin serotype E (rBoNTE(H(c)): antigen E) by Pichia pastoris. Publishing Authors By Initials

J Sinha,M Inan,S Fanders,S Taoka,M Gouthro,T Swanson,R Barent,A Barthuli,BM Loveless,LA Smith,T Smith,I Henderson,J Ross,MM Meagher,

For similar proteins: recombinant proteins research abstracts see: proteins: recombinant proteins research

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Cell bank characterization and fermentation optimization for production of recombinant heavy chain C-terminal fragment of botulinum neurotoxin serotype E (rBoNTE(H(c)): antigen E) by Pichia pastoris. Journal Published:

PUBLICATION TYPE: Research Support, U.S. Gov't,

Journal: Journal of biotechnology

VOLUME: 127

Page Numbers: 462-74

Journal Abbreviation: J. Biotechnol.

ISSN: 0168-1656

DAY: 31

MONTH: 07

YEAR: 2006

Cell bank characterization and fermentation optimization for production of recombinant heavy chain C-terminal fragment of botulinum neurotoxin serotype E (rBoNTE(H(c)): antigen E) by Pichia pastoris. Information

Number of References:

LANGUAGE: eng

NlmUniqueID: 8411927

Cell bank characterization and fermentation optimization for production of recombinant heavy chain C-terminal fragment of botulinum neurotoxin serotype E (rBoNTE(H(c)): antigen E) by Pichia pastoris. Keywords Mesh Terms:

KEYWORDS: Recombinant Proteins

MESH TERMS: isolation & purification

Chemical & Substance for Abstract: Cell bank characterization and fermentation optimization for production of recombinant heavy chain C-terminal fragment of botulinum neurotoxin serotype E (rBoNTE(H(c)): antigen E) by Pichia pastoris. Information

Substance Name: botulinum toxin type E

Registry Number: 0

Grant and Affiliation Information for Cell bank characterization and fermentation optimization for production of recombinant heavy chain C-terminal fragment of botulinum neurotoxin serotype E (rBoNTE(H(c)): antigen E) by Pichia pastoris.

AFFILIATION: Biological Process Development Facility, Department of Chemical and Biomolecular Engineering, University of Nebraska-Lincoln, Lincoln, NE 68588-0466, USA.

Country: Netherlands

AGENCY: United States NIAID

GRANT: U01 AI056514-01

ACRONYM: AI

MEDLINETA: J Biotechnol

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