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J. Bacteriol. doi:10.1128/JB.02008-07
Copyright (c) 2008, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

The sim Operon Facilitates the Transport and Metabolism of Sucrose Isomers in Lactobacillus casei ATCC 334

Microbial Biochemistry and Genetics Unit, Oral Infection and Immunity Branch, NIDCR, National Institutes of Health, DHHS, Bethesda, Maryland 20892; Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, Maryland 20892; and the Proteome Exploration Laboratory, California Institute of Technology, Pasadena, California, 91125

^* To whom correspondence should be addressed. Email: jthompson{at}dir.nidcr.nih.gov.

	Abstract

Inspection of the genome sequence of Lactobacillus casei ATCC 334 revealed two operons with the potential for dissimilation of the five isomers of sucrose. To test this hypothesis, cells of L. casei 334 were grown in a defined medium supplemented with various sugars, including each of the five isomeric disaccharides. Extracts prepared from cells grown on the sucrose isomers contained high levels of two polypeptides of M_r 50 and 17.5 kDa, respectively. Neither protein was present in cells grown on glucose, maltose or sucrose. Proteomic, enzymatic, and Western blot analyses identified the 50 kDa protein as an NAD⁺ and metal ion- dependent phospho- - glucosidase (Pagl). The oligomeric enzyme has been purified and a catalytic mechanism proposed. The smaller polypeptide represented an EIIA component of the phosphoenolpyruvate – dependent: sugar phosphotransferase system (PEP-PTS). Phospho -- glucosidase and EIIA are encoded by genes at loci LSEI_0369 (simA) and LSEI_0374 (simF) respectively, in a block of seven genes comprising the sucrose isomer metabolism (sim) operon. Northern blot analyses provided evidence for the up-regulation of three mRNA transcripts during logarithmic growth of L. casei 334 on sucrose isomers. Internal simA and simF gene probes hybridized to transcripts of 1.5 and 1.3 kb, respectively. A 6.8 kb mRNA transcript was detected by both probes, indicative of cotranscription of the entire sim operon.