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Microbiology 151 (2005), 439-446; DOI  10.1099/mic.0.27411-0
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Microbiology 151 (2005), 439-446; DOI  10.1099/mic.0.27411-0
© 2005 Society for General Microbiology

Molecular characterization of the CmbR activator-binding site in the metC–cysK promoter region in Lactococcus lactis

Natasa Golic1,2,3, Martijn Schliekelmann1,2, María Fernández1,2,{ddagger}, Michiel Kleerebezem1,2 and Richard van Kranenburg1,2,{dagger}

1 Wageningen Centre for Food Sciences, Wageningen, The Netherlands
2 Flavour, Nutrition and Ingredients Department, NIZO Food Research, PO Box 20, 6710 BA Ede, The Netherlands
3 Institute of Molecular Genetics and Genetic Engineering, Belgrade, Yugoslavia

Correspondence
Michiel Kleerebezem
michiel.kleerebezem{at}nizo.nl

The metC–cysK operon involved in sulphur metabolism in Lactococcus lactis is positively regulated by the LysR-type protein CmbR. Transcription from the metC promoter is activated when concentrations of methionine and cysteine in the growth medium are low. The metC promoter region contains two direct and three inverted repeats. Deletion analysis indicated that direct repeat 2 (DR2) is required for activation of the metC promoter by CmbR. Gel mobility shift assays confirmed that CmbR binds to a 407 bp DNA fragment containing the metC promoter. This binding was stimulated by O-acetyl-L-serine. Competition experiments with deletion variants of the metC promoter showed that CmbR binding only occurred with fragments containing an intact DR2, confirming that DR2 is the CmbR binding site within the metC promoter.

Abbreviations: CFE, cell-free extract; DR, direct repeat; LTTR, LysR-type transcriptional regulator; OAS, O-acetyl-L-serine

{dagger}Present address: Purac Biochem, Gorinchem, The Netherlands.

{ddagger}Present address: Instituto de Productos Lácteos de Asturias (CSIC) C/Infiesto s/n Villaviciosa, Asturias, Spain.




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