Sinorhizobium meliloti Flavin Secretion and Bacteria-Host Interaction: Role of the Bifunctional RibBA Protein

Abstract

Sinorhizobium meliloti, the nitrogen-fixing bacterial symbiont of Medicago spp. and other legumes, secretes a considerable amount of riboflavin. This precursor of the cofactors flavin mononucleotide and flavin adenine dinucleotide is a bioactive molecule that has a beneficial effect on plant growth. The ribBA gene of S. meliloti codes for a putative bifunctional enzyme with dihydroxybutanone phosphate synthase and guanosine triphosphate (GTP) cyclohydrolase II activities, catalyzing the initial steps of the riboflavin biosynthesis pathway. We show here that an in-frame deletion of ribBA does not cause riboflavin auxotrophy or affect the ability of S. meliloti to establish an effective symbiosis with the host plant but does affect the ability of the bacteria to secrete flavins, colonize host-plant roots, and compete for nodulation. A strain missing the RibBA protein retains considerable GTP cyclohydrolase II activity. Based on these results, we hypothesize that S. meliloti has two partly interchangeable modules for biosynthesis of riboflavin, one fulfilling the internal need for flavins in bacterial metabolism and the other producing riboflavin for secretion. Our data also indicate that bacteria-derived flavins play a role in communication between rhizobia and the legume host and that the RibBA protein is important in this communication process even though it is not essential for riboflavin biosynthesis and symbiosis.

Laser‐ablation electrospray ionization mass spectrometry with ion mobility separation reveals metabolites in the symbiotic interactions of soybean roots and rhizobia

“…The second highest pathway coverage was the riboflavin metabolism (two of 10 molecules detected), which is biosynthesized in plants and bacteria (Bacher et al ., ) as a direct precursor of the flavin cofactors (McCormick, ). Root‐colonizing bacteria secrete riboflavin as a significant ecological factor in host‐plant root colonization and communication by rhizobia, root respiration and nodulation, and in host‐plant shoot growth (Schwinghamer, ; Yang et al ., ; Yurgel et al ., ). Interestingly, we obtained a pathway coverage of 8% purine metabolism (five of 61 species observed), which is involved in BNF.…”

Comparative gene expression analysis of Porphyromonas gingivalis ATCC 33277 in planktonic and biofilms states

“…aeruginosa [63]. Also, related to quorum sensing signaling, the up regulated gene PGN_0643, has been involved in the biosynthesis of riboflavin, a substance associated in a number of extracellular processes by bacteria, especially Gram-negative organisms [64–66]. …”

A Novel Mechanism for Nitrosative Stress Tolerance Dependent on GTP Cyclohydrolase II Activity Involved in Riboflavin Synthesis of Yeast

“…SDS-PAGE and CBB staining were performed to observe the protein purity. The GTPCH2 activity of the purified Rib1 was measured by monitoring the fluorescence derived from 6,7-dimethylpterin, which is a reaction product of DARP and diacetyl, using a spectrofluorometer F-7000 (Hitachi), as previously described 38,39 . To quantify generated DARP, the standard curve was drawn using various concentrations (0-50 μM) of 6,7-dimethylpterin.…”

“…The GTPCH2 reaction of rRib1p was performed in the solution containing 100 mM Tris-HCl (pH 7.4), 5 mM MgCl 2 , 5 mM DTT, 50 μM GTP, and rRib1p by incubation at 37 °C for 2 h. Enzyme was inactivated by boiling for 5 min and the supernatant after centrifugation was used for further analyses as a DARP containing solution. The concentration of DARP contained in the sample was measured using diacetyl and a spectrofluorometer F-7000 (Hitachi) as described above and previously 38,39 . (2020) 10:6015 | https://doi.org/10.1038/s41598-020-62890-3 www.nature.com/scientificreports www.nature.com/scientificreports/ In vitro NO quenching assay.…”