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Journal of Bacteriology, January 2004, p. 473-480, Vol. 186, No. 2
0021-9193/04/$08.00+0     DOI: 10.1128/JB.186.2.473-480.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

The Sorbitol Phosphotransferase System Is Responsible for Transport of 2-C-Methyl-D-Erythritol into Salmonella enterica Serovar Typhimurium

Charles A. Testa,{dagger} Rita M. Cornish, and C. Dale Poulter*

Department of Chemistry, University of Utah, Salt Lake City, Utah 84112

Received 23 June 2003/ Accepted 8 October 2003

2-C-methyl-D-erythritol 4-phosphate is the first committed intermediate in the biosynthesis of the isoprenoid precursors isopentenyl diphosphate and dimethylallyl diphosphate. Supplementation of the growth medium with 2-C-methyl-D-erythritol has been shown to complement disruptions in the Escherichia coli gene for 1-deoxy-D-xylulose 5-phosphate synthase, the enzyme that synthesizes the immediate precursor of 2-C-methyl-D-erythritol 4-phosphate. In order to be utilized in isoprenoid biosynthesis, 2-C-methyl-D-erythritol must be phosphorylated. We describe the construction of Salmonella enterica serovar Typhimurium strain RMC26, in which the essential gene encoding 1-deoxy-D-xylulose 5-phosphate synthase has been disrupted by insertion of a synthetic mevalonate operon consisting of the yeast ERG8, ERG12, and ERG19 genes, responsible for converting mevalonate to isopentenyl diphosphate under the control of an arabinose-inducible promoter. Random mutagenesis of RMC26 produced defects in the sorbitol phosphotransferase system that prevented the transport of 2-C-methyl-D-erythritol into the cell. RMC26 and mutant strains of RMC26 unable to grow on 2-C-methyl-D-erythritol were incubated in buffer containing mevalonate and deuterium-labeled 2-C-methyl-D-erythritol. Ubiquinone-8 was isolated from these cells and analyzed for deuterium content. Efficient incorporation of deuterium was observed for RMC26. However, there was no evidence of deuterium incorporation into the isoprenoid side chain of ubiquinone Q8 in the RMC26 mutants.

* Corresponding author. Mailing address: Department of Chemistry, University of Utah, Salt Lake City, UT 84112. Phone: (801) 581-6685. Fax: (801) 581-4391. E-mail: poulter{at}chem.utah.edu.

{dagger} Present address: Echelon Biosciences, Inc., Salt Lake City, UT 84108.

Journal of Bacteriology, January 2004, p. 473-480, Vol. 186, No. 2
0021-9193/04/$08.00+0     DOI: 10.1128/JB.186.2.473-480.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



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