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Journal of Bacteriology, February 2003, p. 870-878, Vol. 185, No. 3
0021-9193/03/$08.00+0     DOI: 10.1128/JB.185.3.870-878.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Characterization, Expression, and Mutation of the Lactococcus lactis galPMKTE Genes, Involved in Galactose Utilization via the Leloir Pathway

Benoît P. Grossiord,1,2* Evert J. Luesink,1 Elaine E. Vaughan,3 Alain Arnaud,2 and Willem M. de Vos3

NIZO Food Research, 6710 BA Ede,1 Laboratory of Microbiology, Wageningen University, 6703 CT Wageningen, The Netherlands,3 U.F.R de Microbiologie Industrielle et de Génétique des Microorganismes, Ecole Nationale Supérieure Agronomique, 34060 Montpellier Cédex 1, France2

Received 5 July 2002/ Accepted 6 November 2002

A cluster containing five similarly oriented genes involved in the metabolism of galactose via the Leloir pathway in Lactococcus lactis subsp. cremoris MG1363 was cloned and characterized. The order of the genes is galPMKTE, and these genes encode a galactose permease (GalP), an aldose 1-epimerase (GalM), a galactokinase (GalK), a hexose-1-phosphate uridylyltransferase (GalT), and a UDP-glucose 4-epimerase (GalE), respectively. This genetic organization reflects the order of the metabolic conversions during galactose utilization via the Leloir pathway. The functionality of the galP, galK, galT, and galE genes was shown by complementation studies performed with both Escherichia coli and L. lactis mutants. The GalP permease is a new member of the galactoside-pentose-hexuronide family of transporters. The capacity of GalP to transport galactose was demonstrated by using galP disruption mutant strains of L. lactis MG1363. A galK deletion was constructed by replacement recombination, and the mutant strain was not able to ferment galactose. Disruption of the galE gene resulted in a deficiency in cell separation along with the appearance of a long-chain phenotype when cells were grown on glucose as the sole carbon source. Recovery of the wild-type phenotype for the galE mutant was obtained either by genetic complementation or by addition of galactose to the growth medium.

* Corresponding author. Present address: Laboratoire de Microbiologie et de Biochimie Appliquée, ENITA de Bordeaux, 1 cours du Général de Gaulle, BP201, F-33175 Gradignan Cedex, France, Phone: (33) 557350735. Fax: (33) 557350739. E-mail: b-grossiord{at}enitab.fr.

Journal of Bacteriology, February 2003, p. 870-878, Vol. 185, No. 3
0021-9193/03/$08.00+0     DOI: 10.1128/JB.185.3.870-878.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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