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Contribution of the phosphoenolpyruvate:mannose phosphotransferase system to carbon catabolite repression in Lactobacillus pentosus

EC Slater Institute, BioCentrum, University of Amsterdam, Plantage Muidergracht 12, 1018 TV Amsterdam, The Netherlands¹
TNO Voeding, Department of Applied Microbiology and Gene Technology, PO box 360 3700 AJ Zeist, The Netherlands²

Author for correspondence: Peter H. Pouwels (TNO Voeding). Tel: +31 30 6944 924. Fax: +31 30 6944 466. e-mail: Pouwels{at}voeding.tno.nl

The role of the Lactobacillus pentosus phosphoenolpyruvate:mannose phosphotransferase system (mannose PTS) in sugar transport and control of sugar utilization was investigated. Growth experiments and measurements of PEP-dependent phosphorylation of sugars, of sugar transport and of catabolic enzyme activity were performed, to compare a wild-type strain with an EIIB^Man mutant, LPE6, and a ccpA mutant, LPE4. Fructose uptake in wild-type bacteria demonstrated the presence of two fructose-specific PTSs: a high-affinity system, EII^Fru (K_m=52 µM) which is inducible by fructose, and a low-affinity system (K_m=300 µM). The latter system was lacking in LPE6 and therefore corresponds to EII^Man. LPE6 was unable to phosphorylate glucose, mannose, N-acetylglucosamine and 2-deoxyglucose in a PEP-dependent reaction, indicating that these sugars are substrates of EII^Man. Transport and phosphorylation of these compounds was the same in LPE4 and in wild-type bacteria, although growth of LPE4 on these sugars was impaired. In wild-type bacteria and in LPE4 the activity of EII^Fru was lowered by the presence of EII^Man substrates in the growth medium, but this decrease was not observed in LPE6. These results indicate that EII^Man but not CcpA regulates the synthesis of EII^Fru. Mutations in EII^Man or CcpA resulted in a relief of catabolite repression exerted by EII^Man substrates on the activity of ß-galactosidase and ß-glucosidase, indicating that EII^Man and CcpA are important components in catabolite repression in L. pentosus. Fructose-mediated repression of these two enzymes appeared to be correlated with the activity of EII^Fru.

Keywords: mannose PTS, fructose PTS, CcpA, sugar transport, regulation

Abbreviations: 2DG, 2-deoxyglucose; CR, catabolite repression; mannose PTS, phosphoenolpyruvate:mannose phosphotransferase system; PEP, phosphoenolpyruvate

^a Present address: Institut National de la Recherche Agronomique, Unité Flore Lactique et Environnement Carné, CRJ, Domaine de Vilvert, 78350 Jouy-en-Josas, France.

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