Novel mixed-linkage β-glucan activated by c-di-GMP in Sinorhizobium meliloti

An artificial increase of cyclic diguanylate (c-di-GMP) levels in Sinorhizobium meliloti 8530, a bacterium that does not carry known cellulose synthesis genes, leads to overproduction of a substance that binds the dyes Congo red and calcofluor. Sugar composition and methylation analyses and NMR studies identified this compound as a linear mixed-linkage (1→3)(1→4)-β-D-glucan (ML β-glucan), not previously described in bacteria but resembling ML β-glucans found in plants and lichens. This unique polymer is hydrolyzed by the specific endoglucanase lichenase, but, unlike lichenan and barley glucan, it generates a disaccharidic →4)-β-DGlcp-(1→3)-β-D-Glcp-(1→ repeating unit. A two-gene operon bgsBA required for production of this ML β-glucan is conserved among several genera within the order Rhizobiales, where bgsA encodes a glycosyl transferase with domain resemblance and phylogenetic relationship to curdlan synthases and to bacterial cellulose synthases. ML β-glucan synthesis is subjected to both transcriptional and posttranslational regulation. bgsBA transcription is dependent on the exopolysaccharide/quorum sensing ExpR/SinI regulatory system, and posttranslational regulation seems to involve allosteric activation of the ML β-glucan synthase BgsA by c-di-GMP binding to its C-terminal domain. To our knowledge, this is the first report on a linear mixed-linkage (1→3)(1→4)-β-glucan produced by a bacterium. The S. meliloti ML β-glucan participates in bacterial aggregation and biofilm formation and is required for efficient attachment to the roots of a host plant, resembling the biological role of cellulose in other bacteria.Ministerio de Economía y Competittividad BIO2011-23032Junta de Andalucía P10-CVI-5800Consejo Superior de Investigaciones Científicas 201440E02

Novel mixed-linkage β-glucan activated by c-di-GMP in Sinorhizobium meliloti

© 2015 PNAS. An artificial increase of cyclic diguanylate (c-di-GMP) levels in Sinorhizobium meliloti 8530, a bacterium that does not carry known cellulose synthesis genes, leads to overproduction of a substance that binds the dyes Congo red and calcofluor. Sugar composition and methylation analyses and NMR studies identified this compound as a linear mixed-linkage (1→3)(1→4)-β-D-glucan (ML β-glucan), not previously described in bacteria but resembling ML β-glucans found in plants and lichens. This unique polymer is hydrolyzed by the specific endoglucanase lichenase, but, unlike lichenan and barley glucan, it generates a disaccharidic →4)-β-DGlcp-(1→3)-β-D-Glcp-(1→ repeating unit. A two-gene operon bgsBA required for production of this ML β-glucan is conserved among several genera within the order Rhizobiales, where bgsA encodes a glycosyl transferase with domain resemblance and phylogenetic relationship to curdlan synthases and to bacterial cellulose synthases. ML β-glucan synthesis is subjected to both transcriptional and posttranslational regulation. bgsBA transcription is dependent on the exopolysaccharide/quorum sensing ExpR/SinI regulatory system, and posttranslational regulation seems to involve allosteric activation of the ML β-glucan synthase BgsA by c-di-GMP binding to its C-terminal domain. To our knowledge, this is the first report on a linear mixed-linkage (1→3)(1→4)-β-glucan produced by a bacterium. The S. meliloti ML β-glucan participates in bacterial aggregation and biofilm formation and is required for efficient attachment to the roots of a host plant, resembling the biological role of cellulose in other bacteria.We thank J. Nogales for Sme expR and sinI derivatives; M. J. Pérez-Mendoza, University of Granada, and I. Rodríguez-García, University of Almería, for help with initial chemical analyses; T. Felipe-Reyes and D. Rodríguez-Carbonell (Estación Experimental del Zaidín, EEZ) for excellent technical assistance; S. Muñoz for help with competitive nodulation; A. Olmedilla, EEZ, and G. Martín, University of Malaga, for help with scanning microscopy; J. Düvel, Helmholtz Centre for Infection Research, for assistance with dot blot assays and for providing cloned PA3353; and Centro de Investigación Tecnología e Innovación de la Universidad de Sevilla for support with NMR studies. This work was supported by Grant BIO2011-23032 from the Ministerio de Economía y Competitividad and Grant P10-CVI-5800 from the Junta de Andalucía, both cofinanced by the European Fund for Economic and Regional Development, and by Consejo Superior de Investigaciones Cientificas (CSIC) Grant 201440E026. D.P.-M. was supported by a Junta de Ampliación de Estudios (JAE)-doc CSIC contract and later by Grants P10-CVI-5800 and 201440E026; L.R.-J. was supported by a JAE-Pre fellowship; and G.d.A.F. was supported by a contract associated with Grant P10-CVI-5800.Peer Reviewe