Transcriptional and functional characterization of genetic elements involved in galacto-oligosaccharide utilization by Bifidobacterium breve UCC2003.
Bottom Line: We further demonstrate that an extracellular endogalactanase specified by several B. breve strains, including B. breve UCC2003, is essential for partial degradation of PGOS components with a high degree of polymerization.These partially hydrolysed PGOS components are presumed to be transported into the bifidobacterial cell via various ABC transport systems and sugar permeases where they are further degraded to galactose and glucose monomers that feed into the bifid shunt.This work significantly advances our molecular understanding of bifidobacterial PGOS metabolism and its associated genetic machinery to utilize this prebiotic.
Affiliation: Alimentary Pharmabiotic Centre, National University of Ireland, Western Road, Cork, Ireland; Departments of Microbiology, National University of Ireland, Western Road, Cork, Ireland.Show MeSH
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Mentions: In order to investigate which genes may be involved in PGOS metabolism in B. breve UCC2003, global gene expression was determined by microarray analysis during growth of this bifidobacterial strain on PGOS and this was compared with its expression pattern when grown on ribose as sole carbohydrate source. Total RNA was isolated from mid-log phase cultures of B. breve UCC2003 grown on PGOS or ribose as the sole carbohydrate source. Analysis of the DNA microarray data revealed that the expression of four gene clusters was significantly upregulated when UCC2003 was grown on PGOS (fold change > 3.0, P < 0.001; Table 1 and Fig. 3). The first GOS-inducible cluster of six genes, galC, galD, galE galG, galR and galA, constitute the previously identified galactan metabolism cluster (O'Connell Motherway et al., 2011b), while the second cluster, Bbr_1551 to Bbr_1553, consists of three genes specifying a predicted galactoside symporter and a GH2 family β-galactosidase in divergent orientations, designated here as lacS and lacZ respectively, and an associated gene encoding a putative LacI-type transcriptional regulator. The third GOS-inducible cluster comprises five genes, Bbr_0526 to Bbr_530, that are predicted to specify a LacI-type transcriptional regulator, two ABC-type permeases, a GH42 family β-galactosidase and a solute-binding protein, which are designated gosR, gosD, gosE gosG and gosC respectively. The fourth set of upregulated genes are the presumed galT and galK genes that encode Galactose-1-phosphate uridyltransferase and galactokinase respectively (Kitaoka et al., 2005; Fushinobu, 2010), and represent key enzymes of the Leloir pathway that converts galactose to d-glucose 6-phosphate that can feed into the bifid shunt (Fushinobu, 2010; Pokusaeva et al., 2011a) (Fig. 3).
Affiliation: Alimentary Pharmabiotic Centre, National University of Ireland, Western Road, Cork, Ireland; Departments of Microbiology, National University of Ireland, Western Road, Cork, Ireland.