Genetic manipulation of acid formation pathways by gene inactivation in Clostridium acetobutylicum ATCC 824.

Integrational plasmid technology has been used to disrupt metabolic pathways leading to acetate and butyrate formation in Clostridium acetobutylicum ATCC 824. Non-replicative plasmid constructs, containing either clostridial phosphotransacetylase (pta) or butyrate kinase (buk) gene fragments, were integrated into homologous regions on the chromosome. Integration was assumed to occur by a Campbell-like mechanism, inactivating either pta or buk. Inactivation of the pta gene reduced phosphotransacetylase and acetate kinase activity and significantly decreased acetate production. Inactivation of the buk gene reduced butyrate kinase activity, significantly decreased butyrate production and increased butanol production.

[1]  M. Young.  Gene amplification in Bacillus subtilis. , 1984, Journal of general microbiology.

[2]  G. Bennett,et al.  Inactivation of an aldehyde/alcohol dehydrogenase gene from Clostridium acetobutylicum ATCC 824. , 1996, Applied biochemistry and biotechnology.

[3]  M. Young,et al.  Recent advances in the genetics of the clostridia. , 1989, FEMS microbiology reviews.

[4]  C. M. Murphy,et al.  Lactococcus and Lactobacillus , 1993 .

[5]  Inactivation of an aldehyde/alcohol dehydrogenase gene fromClostridium acetobutylicum ATCC 824 , 1996 .

[6]  M. Hartmanis.  Butyrate kinase from Clostridium acetobutylicum. , 1987, The Journal of biological chemistry.

[7]  P. Piggot,et al.  Use of integrational plasmid vectors to demonstrate the polycistronic nature of a transcriptional unit (spoIIA) required for sporulation of Bacillus subtilis. , 1984, Journal of general microbiology.

[8]  S. Ehrlich,et al.  Stability of reiterated sequences in the Bacillus subtilis chromosome , 1989, Journal of bacteriology.

[9]  E. Papoutsakis,et al.  Phosphotransbutyrylase from Clostridium acetobutylicum ATCC 824 and its role in acidogenesis , 1989, Applied and environmental microbiology.

[10]  D. T. Jones,et al.  Acetone-butanol fermentation revisited. , 1986, Microbiological reviews.

[11]  S. Ehrlich,et al.  Plasmid replication stimulates DNA recombination in Bacillus subtilis. , 1987, Journal of molecular biology.

[12]  J. Sambrook,et al.  Molecular Cloning: A Laboratory Manual , 2001 .

[13]  H. Kornberg,et al.  The enzymic interconversion of acetate and acetyl-coenzyme A in Escherichia coli. , 1977, Journal of general microbiology.

[14]  G. Bennett,et al.  Intracellular Concentrations of Coenzyme A and Its Derivatives from Clostridium acetobutylicum ATCC 824 and Their Roles in Enzyme Regulation , 1994, Applied and environmental microbiology.

[15]  G. Gottschalk,et al.  Physiological Events in Clostridium acetobutylicum during the Shift from Acidogenesis to Solventogenesis in Continuous Culture and Presentation of a Model for Shift Induction , 1992, Applied and environmental microbiology.

[16]  D. Hopwood,et al.  Genetic manipulation of Streptomyces: integrating vectors and gene replacement. , 1991, Methods in enzymology.

[17]  E. Papoutsakis,et al.  Cloning and expression of Clostridium acetobutylicum phosphotransbutyrylase and butyrate kinase genes in Escherichia coli , 1988, Journal of bacteriology.

[18]  Evaluation of macrolide and lincosamide antibiotics for plasmid maintenance in low pH Clostridium acetobutylicum ATCC 824 fermentations , 1993 .

[19]  I. A. Rose.  [97] Acetate kinase of bacteria (acetokinase): Acetate + ATP ⇄ Acetyl-P + ADP☆ , 1955 .

[20]  E. Simon.  The formation of lactic acid by Clostridium acetobutylicum (Weizmann). , 1947, Archives of biochemistry.

[21]  E. Papoutsakis,et al.  Cloning, structure, and expression of acid and solvent pathway genes of Clostridium acetobutylicum. , 1993, Biotechnology.

[22]  G. Bennett,et al.  Sequence and arrangement of genes encoding sigma factors in Clostridium acetobutylicum ATCC 824. , 1995, Gene.

[23]  E. Papoutsakis,et al.  The effect of pH on nitrogen supply, cell lysis, and solvent production in fermentations of Clostridium acetobutylicum , 1985, Biotechnology and bioengineering.

[24]  E. Papoutsakis,et al.  Molecular characterization of an aldehyde/alcohol dehydrogenase gene from Clostridium acetobutylicum ATCC 824 , 1994, Journal of bacteriology.

[25]  G. Venema,et al.  Campbell-like integration of heterologous plasmid DNA into the chromosome of Lactococcus lactis subsp. lactis , 1989, Applied and environmental microbiology.

[26]  E. Papoutsakis,et al.  Molecular characterization of two Clostridium acetobutylicum ATCC 824 butanol dehydrogenase isozyme genes , 1992, Journal of bacteriology.

[27]  E. Southern.  Detection of specific sequences among DNA fragments separated by gel electrophoresis. , 1975, Journal of molecular biology.