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Overexpression of the ftsZ gene from Corynebacterium glutamicum (Brevibacterium lactofermentum) in Escherichia coli

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Abstract:

Our goal in this work was to overexpress the essential cell division FtsZ protein from Corynebacterium glutamicum (Brevibacterium lactofermentum) (FtsZCG) in Escherichia coli to produce anti-FtsZCG polyclonal antibodies. Previous results from our laboratory showed that ftsZCG was not expressed in E. coli in a sufficient amount to purify FtsZCG. However, when ftsZCG (without upstream sequences) was transcriptionally fused to the T7 promoter, different truncated FtsZCG proteins (28–32 kDa) were overexpressed in E. coli, and in all cases, stop codons were created because of DNA deletions or rearrangements. Nevertheless, we were able to overexpress and purify an N-terminally hexa-His-tagged FtsZCG from uninduced E. coli cells carrying a pET-28a(+) derivative, yielding about 5 mg of 98% pure protein per 100-mL culture.Key words: Brevibacterium lactofermentum, Corynebacterium glutamicum, FtsZ overexpresssion, hexa-His-tagged FtsZ.

Le but de ces travaux était de surexprimer la protéine essentielle à la division cellulaire FtsZ de Corynebacterium glutamicum (Brevibacterium lactofermentum) (FtsZCG) chez Escherichia coli afin de produire des anticorps polyclonaux anti-FtsZCG. Les résultats antérieurs de notre laboratoire ont montré que ftsZCG n'était pas exprimé chez E. coli en quantités suffisantes pour purifier FtsZCG. Toutefois, lorsque ftsZCG (sans les séquences en amont) fut fusionné transcriptionnellement au promoteur T7, diverses protéines FtsZCG tronquées (28–32 kDa) ont été surexprimées chez E. coli, et dans tous les cas des codons de terminaison ont été introduits par des délétions ou des réarrangements d'ADN. Néanmoins, nous avons été capables de surexprimer et de purifier de la FtsZCG pourvue d'une étiquette N-terminale hexa-His de cellules de E. coli non induites contenant un dérivé de pET-28a(+), permettant la production d'environ 5 mg de protéines pures à 98 % par 100 mL de culture.Mots clés : Brevibacterium lactofermentum, Corynebacterium glutamicum, surexpression de FtsZ, FtsZCG avec étiquette hexa-His.[Traduit par la Rédaction]

Document Type: Research Article

Publication date: January 1, 2005

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  • Published since 1954, this monthly journal contains new research in the field of microbiology including applied microbiology and biotechnology; microbial structure and function; fungi and other eucaryotic protists; infection and immunity; microbial ecology; physiology, metabolism and enzymology; and virology, genetics, and molecular biology. It also publishes review articles and notes on an occasional basis, contributed by recognized scientists worldwide.
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