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GENETICS, GENOMICS, AND MOLECULAR EVOLUTION

Characterization of Mannuronan C-5-Epimerase Genes from the Brown Alga Laminaria digitata¹

Unité Mixte de Recherche 1931, Centre National de la Recherche Scientifique and Laboratoires Goëmar, Station Biologique de Roscoff, BP 74, 29682 Roscoff cedex, Brittany, France

Alginate is an industrially important polysaccharide obtained commercially by harvesting brown algae. The final step in alginate biosynthesis, the epimerization of -1,4-D-mannuronic acid to -1,4-L-guluronic acid, a structural change that controls the physicochemical properties of the alginate, is catalyzed by the enzyme mannuronan C-5-epimerase. Six different cDNAs with homology to bacterial mannuronan C-5-epimerases were isolated from the brown alga Laminaria digitata (Phaeophyceae). Hydrophobic cluster analysis indicated that the proteins encoded by the L. digitata sequences have important structural similarities to the bacterial mannuronan C-5-epimerases, including conservation of the catalytic site. The expression of the C-5-epimerase genes was examined by northern-blot analysis and reverse transcriptase-polymerase chain reaction in L. digitata throughout a year. Expression was also monitored in protoplast cultures by northern and western blot, reverse transcriptase-polymerase chain reaction, and activity measurements. From both the structural comparisons and the expression pattern, it appears that the cDNAs isolated from L. digitata encode functional mannuronan C-5-epimerases. The phylogenetic relationships of the bacterial and brown algal enzymes and the inferences on the origin of alginate biosynthetic machinery are discussed.

¹ This work was supported by the European Union Grant Polyeng (QLK3-CT-1999–00034).

^* Corresponding author; e-mail: boyen{at}sb-roscoff.fr; fax 33–0298292324.

Received April 25, 2003; returned for revision June 9, 2003; accepted July 11, 2003.

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