Thermoanaerobacter brockii alcohol dehydrogenase promoter...

Chemistry: molecular biology and microbiology – Micro-organism – tissue cell culture or enzyme using process... – Recombinant dna technique included in method of making a...

Reexamination Certificate

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C435S320100, C435S254110, C435S252300, C536S024100

Reexamination Certificate

active

10389821

ABSTRACT:
The present invention discloses the isolation and use of a specific bacterial promoter region suitable for use in constructs for the high level production of heterologous proteins. This promoter is derived from the bacterial gene encoding for alcohol dehydrogenase, in particular the alcohol dehydrogenase genes isolated from the thermophilic bacterial strainT. brockiiand the mesophilic bacterial strainClostridium beijerinckii. It is now disclosed that using either the intact promoter region or certain specific fragments consisting of at least a 88 bp DNA sequence in the upstream untranslated region of the bacterial alcohol dehydrogenase gene, operatively linked to the nucleic acid sequences encoding a heterologous protein, and insertion into a DNA plasmid or any other suitable vector system, heterologous genes can be expressed in high levels in host cells. Heterologous proteins or peptides can be expressed constitutively at high levels. The proteins are obtained in their active folded form.

REFERENCES:
patent: 5908924 (1999-06-01), Burdette et al.
Khlebnikov et al. Modulation of gene expression from the arabinose-inducible araBAD promoter. J Ind Microbiol Biotechnol. vol. 29, No. 1, pp. 34-37, Jul. 2002.
GenBank Accession No. AF157307.1, publicly available Sep. 14, 1999.
Peretz et al. Molecular cloning, nucleotide sequenceing, and expression of genes encoding alcohol dehydrogenases from the thermophile Thermoanaerobacter brockii and the mesophile Clostridium beijerinckii. Anaerobe, vol. 3, pp. 259-270, 1997.
McDonald et al. Large-scale purification and characterization of recombinant fibroblast growth factor-saporin mitotoxin. Protein Expression and Purification, vol. 8, pp. 97-108, 1996.
Burdette et al. Cloning and expression of the gene ecoding the Thermoanaerobacter ethanolicus 39E secondar-alcohol dehydrogenase and biochemical characterization of the enzyme. Biochem J., vol. 316, pp. 115-122, 1996.

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