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Applied and Environmental Microbiology, August 2003, p. 4951-4965, Vol. 69, No. 8
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.8.4951-4965.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Overexpression of groESL in Clostridium acetobutylicum Results in Increased Solvent Production and Tolerance, Prolonged Metabolism, and Changes in the Cell's Transcriptional Program

Christopher A. Tomas,1 Neil E. Welker,2 and Eleftherios T. Papoutsakis1*

Department of Chemical Engineering,1 Department of Biochemistry, Molecular Biology, and Cell Biology, Northwestern University, Evanston, Illinois 602082

Received 24 February 2003/ Accepted 2 June 2003

DNA array and Western analyses were used to examine the effects of groESL overexpression and host-plasmid interactions on solvent production in Clostridium acetobutylicum ATCC 824. Strain 824(pGROE1) was created to overexpress the groESL operon genes from a clostridial thiolase promoter. The growth of 824(pGROE1) was inhibited up to 85% less by a butanol challenge than that of the control strain, 824(pSOS95del). Overexpression of groESL resulted in increased final solvent titers 40% and 33% higher than those of the wild type and plasmid control strains, respectively. Active metabolism lasted two and one half times longer in 824(pGROE1) than in the wild type. Transcriptional analysis of 824(pGROE1) revealed increased expression of motility and chemotaxis genes and a decrease in the expression of the other major stress response genes. Decreased expression of the dnaKJ operon upon overexpression of groESL suggests that groESL functions as a modulator of the CIRCE regulon, which is shown here to include the hsp90 gene. Analysis of the plasmid control strain 824(pSOS95del) revealed complex host-plasmid interactions relative to the wild-type strain, resulting in prolonged biphasic growth and metabolism. Decreased expression of four DNA gyrases resulted in differential expression of many key primary metabolism genes. The ftsA and ftsZ genes were expressed at higher levels in 824(pSOS95del), revealing an altered cell division and sporulation pattern. Both transcriptional and Western analyses revealed elevated stress protein expression in the plasmid-carrying strain.

* Corresponding author. Mailing address: Department of Chemical Engineering, Northwestern University, Evanston, IL 60208. Phone: (847) 491-7455. Fax: (847) 491-3728. E-mail: e-paps{at}northwestern.edu.

Applied and Environmental Microbiology, August 2003, p. 4951-4965, Vol. 69, No. 8
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.8.4951-4965.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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