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J. Biol. Chem., Vol. 281, Issue 48, 36864-36873, December 1, 2006

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The -Phosphoglucomutase of Lactococcus lactis Is Unrelated to the -D-Phosphohexomutase Superfamily and Is Encoded by the Essential Gene pgmH^*

Ana R. Neves¹, Wietske A. Pool, Rute Castro, Ana Mingote, Filipe Santos, Jan Kok, Oscar P. Kuipers, and Helena Santos²

From the Instituto de Tecnologia Química e Biológica and Instituto de Biologia Experimental e Tecnológica, Universidade Nova de Lisboa, Rua da Quinta Grande, 6, Apartado 127, 2780-156 Oeiras, Portugal and the Department of Molecular Genetics, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, 9750 AA Haren, The Netherlands

-Phosphoglucomutase (-PGM) plays an important role in carbohydrate metabolism by catalyzing the reversible conversion of -glucose 1-phosphate to glucose 6-phosphate. Isolation of -PGM activity from cell extracts of Lactococcus lactis strain MG1363 led to the conclusion that this activity is encoded by yfgH, herein renamed pgmH. Its gene product has no sequence homology to proteins in the -D-phosphohexomutase superfamily and is instead related to the eukaryotic phosphomannomutases within the haloacid dehalogenase superfamily. In contrast to known bacterial -PGMs, this 28-kDa enzyme is highly specific for -glucose 1-phosphate and glucose 6-phosphate and showed no activity for mannose phosphate. To elucidate the function of pgmH, the metabolism of glucose and galactose was characterized in mutants overproducing or with a deficiency of -PGM activity. Overproduction of -PGM led to increased glycolytic flux and growth rate on galactose. Despite several attempts, we failed to obtain a deletion mutant of pgmH. The essentiality of this gene was proven by using a conditional knock-out strain in which a native copy of the gene was provided in trans under the control of the nisin promoter. Growth of this strain was severely impaired when -PGM activity was below the control level. We show that the novel L. lactis -PGM is the only enzyme that mediates the interconversion of -glucose 1-phosphate to glucose 6-phosphate and is essential for growth.

Received for publication, July 25, 2006 , and in revised form, September 14, 2006.

^* This work was supported in part by Contract QLK1-CT-2000-01376 from the Commission of the European Communities and Contract POCTI/BIO/48333/2002 from the Fundação para a Ciência e a Tecnologia. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

The on-line version of this article (available at http://www.jbc.org) contains supplemental Figs. 1SM-3SM and Tables 1SM and 2SM.

¹ Supported by a postdoctoral fellowship from the Fundação para a Ciência e a Tecnologia.

² To whom correspondence should be addressed. Tel.: 351-21-446-9828; Fax: 351-21-442-8766; E-mail: santos{at}itqb.unl.pt.

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