Aufsatz in einer Fachzeitschrift
A novel c-di-GMP binding domain in glycosyltransferase BgsA is responsible for the synthesis of a mixed-linkage beta-glucan
Details zur Publikation
Autor(inn)en: | Perez-Mendoza, D.; Bertinetti, D.; Lorenz, R.; Gallegos, M.; Herberg, F.; Sanjuan, J. |
Publikationsjahr: | 2017 |
Zeitschrift: | Scientific Reports |
Seitenbereich: | 8997 |
Jahrgang/Band : | 7 |
Heftnummer: | 1 |
ISSN: | 2045-2322 |
DOI-Link der Erstveröffentlichung: |
Zusammenfassung, Abstract
BgsA is the glycosyltransferase (GT) involved in the synthesis of a linear mixed-linkage beta-glucan (MLG), a recently described exopolysaccharide activated by c-di-GMP in Sinorhizobium meliloti and other Rhizobiales. Although BgsA displays sequence and structural homology with bacterial cellulose synthases (CS), it does not contain any predictable c-di-GMP binding domain. In this work we demonstrate that the cytoplasmic C-terminal domain of BgsA (C-BgsA) binds c-di-GMP with both high affinity (KD = 0.23 muM) and specificity. C-BgsA is structurally different to the otherwise equivalent cytoplasmic C-terminal domain of CS, and does not contain PilZ motifs for c-di-GMP recognition. A combination of random and site-directed mutagenesis with surface plasmon resonance (SPR) allowed identification of the C-BgsA residues which are important not only for c-di-GMP binding, but also for BgsA GT activity. The results suggest that the C-BgsA domain is important for both, c-di-GMP binding and GT activity of BgsA. In contrast to bacterial CS where c-di-GMP has been proposed as a derepressor of GT activity, we hypothesize that the C-terminal domain of BgsA plays an active role in BgsA GT activity upon binding c-di-GMP.
BgsA is the glycosyltransferase (GT) involved in the synthesis of a linear mixed-linkage beta-glucan (MLG), a recently described exopolysaccharide activated by c-di-GMP in Sinorhizobium meliloti and other Rhizobiales. Although BgsA displays sequence and structural homology with bacterial cellulose synthases (CS), it does not contain any predictable c-di-GMP binding domain. In this work we demonstrate that the cytoplasmic C-terminal domain of BgsA (C-BgsA) binds c-di-GMP with both high affinity (KD = 0.23 muM) and specificity. C-BgsA is structurally different to the otherwise equivalent cytoplasmic C-terminal domain of CS, and does not contain PilZ motifs for c-di-GMP recognition. A combination of random and site-directed mutagenesis with surface plasmon resonance (SPR) allowed identification of the C-BgsA residues which are important not only for c-di-GMP binding, but also for BgsA GT activity. The results suggest that the C-BgsA domain is important for both, c-di-GMP binding and GT activity of BgsA. In contrast to bacterial CS where c-di-GMP has been proposed as a derepressor of GT activity, we hypothesize that the C-terminal domain of BgsA plays an active role in BgsA GT activity upon binding c-di-GMP.
