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Applied and Environmental Microbiology, September 2008, p. 5731-5740, Vol. 74, No. 18
0099-2240/08/$08.00+0     doi:10.1128/AEM.00230-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Regulation of Lactobacillus casei Sorbitol Utilization Genes Requires DNA-Binding Transcriptional Activator GutR and the Conserved Protein GutM

Laboratorio de Bacterias Lácticas y Probióticos, IATA-CSIC, P.O. Box 73, 46100 Burjassot, Valencia, Spain,¹ Laboratorie de Microbiologie et Génétique Moléculaire, INRA-AgroParisTech-CNRS, F-78850 Thiverval-Grignon, France²

Received 25 January 2008/ Accepted 22 July 2008

Sequence analysis of the five genes (gutRMCBA) downstream from the previously described sorbitol-6-phosphate dehydrogenase-encoding Lactobacillus casei gutF gene revealed that they constitute a sorbitol (glucitol) utilization operon. The gutRM genes encode putative regulators, while the gutCBA genes encode the EIIC, EIIBC, and EIIA proteins of a phosphoenolpyruvate-dependent sorbitol phosphotransferase system (PTS^Gut). The gut operon is transcribed as a polycistronic gutFRMCBA messenger, the expression of which is induced by sorbitol and repressed by glucose. gutR encodes a transcriptional regulator with two PTS-regulated domains, a galactitol-specific EIIB-like domain (EIIB^Gat domain) and a mannitol/fructose-specific EIIA-like domain (EIIA^Mtl domain). Its inactivation abolished gut operon transcription and sorbitol uptake, indicating that it acts as a transcriptional activator. In contrast, cells carrying a gutB mutation expressed the gut operon constitutively, but they failed to transport sorbitol, indicating that EIIBC^Gut negatively regulates GutR. A footprint analysis showed that GutR binds to a 35-bp sequence upstream from the gut promoter. A sequence comparison with the presumed promoter region of gut operons from various firmicutes revealed a GutR consensus motif that includes an inverted repeat. The regulation mechanism of the L. casei gut operon is therefore likely to be operative in other firmicutes. Finally, gutM codes for a conserved protein of unknown function present in all sequenced gut operons. A gutM mutant, the first constructed in a firmicute, showed drastically reduced gut operon expression and sorbitol uptake, indicating a regulatory role also for GutM.

Published ahead of print on 1 August 2008.