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Journal of Bacteriology, July 2001, p. 3991-3998, Vol. 183, No. 13
0021-9193/01/$04.00+0   DOI: 10.1128/JB.183.13.3991-3998.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Cell Wall Core Galactofuran Synthesis Is Essential for Growth of Mycobacteria

Fei Pan, Mary Jackson, Yufang Ma, and Michael McNeil^*

Department of Microbiology, Colorado State University, Fort Collins, Colorado 80523

Received 29 November 2000/Accepted 10 April 2001

The mycobacterial cell wall core consists of an outer lipid (mycolic acid) layer attached to peptidoglycan via a galactofuranosyl-containing polysaccharide, arabinogalactan. This structural arrangement strongly suggests that galactofuranosyl residues are essential for the growth and viability of mycobacteria. Galactofuranosyl residues are formed in nature by a ring contraction of UDP-galactopyranose to UDP-galactofuranose catalyzed by the enzyme UDP-galactopyranose mutase (Glf). In Mycobacterium tuberculosis the glf gene overlaps, by 1 nucleotide, a gene, Rv3808c, that has been shown to encode a galactofuranosyl transferase. We demonstrate here that glf can be knocked out in Mycobacterium smegmatis by allelic replacement only in the presence of two rescue plasmids carrying functional copies of glf and Rv3808c. The glf rescue plasmid was designed with a temperature-sensitive origin of replication and the M. smegmatis glf knockout mutant is unable to grow at the higher temperature at which the glf-containing rescue plasmid is lost. In a separate experiment, the Rv3808c rescue plasmid was designed with a temperature-sensitive origin of replication and the glf-bearing plasmid was designed with a normal original of replication; this strain was also unable to grow at the nonpermissive temperature. Thus, both glf and Rv3808c are essential for growth. These findings and the fact that galactofuranosyl residues are not found in humans supports the development of UDP-galactopyranose mutase and galactofuranosyl transferase as important targets for the development of new antituberculosis drugs.

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^* Corresponding author. Mailing address: Department of Microbiology, Colorado State University, Fort Collins, CO 80523. Phone: (970) 491-1784. Fax: (970) 491-1815. E-mail: mmcneil{at}cvmbs.colostate.edu.

Present address: Institut Pasteur, Unité de Génétique Mycobactérienne, 75724 Paris Cedex 15, France.

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Journal of Bacteriology, July 2001, p. 3991-3998, Vol. 183, No. 13
0021-9193/01/$04.00+0   DOI: 10.1128/JB.183.13.3991-3998.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

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