title: Two alternative pathways for the synthesis of the rare compatible solute mannosylglucosylglycerate in Petrotoga mobilis.
abstract: The compatible solute mannosylglucosylglycerate (MGG), recently identified in Petrotoga miotherma, also accumulates in Petrotoga mobilis in response to hyperosmotic conditions and supraoptimal growth temperatures. Two functionally connected genes encoding a glucosyl-3-phosphoglycerate synthase (GpgS) and an unknown glycosyltransferase (gene Pmob_1143), which we functionally characterized as a mannosylglucosyl-3-phosphoglycerate synthase and designated MggA, were identified in the genome of Ptg. mobilis. This enzyme used the product of GpgS, glucosyl-3-phosphoglycerate (GPG), as well as GDP-mannose to produce mannosylglucosyl-3-phosphoglycerate (MGPG), the phosphorylated precursor of MGG. The MGPG dephosphorylation was determined in cell extracts, and the native enzyme was partially purified and characterized. Surprisingly, a gene encoding a putative glucosylglycerate synthase (Ggs) was also identified in the genome of Ptg. mobilis, and an active Ggs capable of producing glucosylglycerate (GG) from ADP-glucose and d-glycerate was detected in cell extracts and the recombinant enzyme was characterized, as well. Since GG has never been identified in this organism nor was it a substrate for the MggA, we anticipated the existence of a nonphosphorylating pathway for MGG synthesis. We putatively identified the corresponding gene, whose product had some sequence homology with MggA, but it was not possible to recombinantly express a functional enzyme from Ptg. mobilis, which we named mannosylglucosylglycerate synthase (MggS). In turn, a homologous gene from Thermotoga maritima was successfully expressed, and the synthesis of MGG was confirmed from GDP-mannose and GG. Based on the measurements of the relevant enzyme activities in cell extracts and on the functional characterization of the key enzymes, we propose two alternative pathways for the synthesis of the rare compatible solute MGG in Ptg. mobilis.
authors.affiliation: Centro de Neurociências e Biologia Celular, Universidade de Coimbra, 3004-517 Coimbra, Portugal.
authors.forename: Chantal
authors.initials: C
authors.lastname: Fernandes
authors.forename: Vitor
authors.initials: V
authors.lastname: Mendes
authors.forename: Joana
authors.initials: J
authors.lastname: Costa
authors.forename: Nuno
authors.initials: N
authors.lastname: Empadinhas
authors.forename: Carla
authors.initials: C
authors.lastname: Jorge
authors.forename: Pedro
authors.initials: P
authors.lastname: Lamosa
authors.forename: Helena
authors.initials: H
authors.lastname: Santos
authors.forename: Milton S
authors.initials: MS
authors.lastname: da Costa
mesh_terms: D001419:Bacteria / Q000187:drug effects / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D002236:Carbohydrate Conformation; D004187:Disaccharides / Q000737:chemistry / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology; D005988:Glyceric Acids / Q000737:chemistry / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D012965:Sodium Chloride / Q000494:pharmacology; D013696:Temperature
chemical_list: D001426:Bacterial Proteins; D004187:Disaccharides; D005988:Glyceric Acids; C522541:mannopyranosyl-1-2-glucopyranosyl-1-2-glycerate; D012965:Sodium Chloride