A unique combination of genetic systems for the synthesis of trehalose in Rubrobacter xylanophilus: properties of a rare actinobacterial TreT.
Trehalose is the primary organic solute in Rubrobacter xylanophilus under all conditions tested, including those for optimal growth. We detected genes of four different pathways for trehalose synthesis in the genome of this organism, namely, the trehalose-6-phosphate synthase (Tps)/trehalose-6-phosphate phosphatase (Tpp), TreS, TreY/TreZ, and TreT pathways. Moreover, R. xylanophilus is the only known member of the phylum Actinobacteria to harbor TreT. The Tps sequence is typically bacterial, but the Tpp sequence is closely related to eukaryotic counterparts. Both the Tps/Tpp and the TreT pathways were active in vivo, while the TreS and the TreY/TreZ pathways were not active under the growth conditions tested and appear not to contribute to the levels of trehalose observed. The genes from the active pathways were functionally expressed in Escherichia coli, and Tps was found to be highly specific for GDP-glucose, a rare feature among these enzymes. The trehalose-6-phosphate formed was specifically dephosphorylated to trehalose by Tpp. The recombinant TreT synthesized trehalose from different nucleoside diphosphate-glucose donors and glucose, but the activity in R. xylanophilus cell extracts was specific for ADP-glucose. The TreT could also catalyze trehalose hydrolysis in the presence of ADP, but with a very high K(m). Here, we functionally characterize two systems for the synthesis of trehalose in R. xylanophilus, a representative of an ancient lineage of the actinobacteria, and discuss a possible scenario for the exceptional occurrence of treT in this extremophilic bacterium.
Nobre
Ana
A
Centro de Neurociências e Biologia Celular, Departamento de Zoologia, Universidade de Coimbra, 3004-517 Coimbra, Portugal.
Alarico
Susana
S
Fernandes
Chantal
C
Empadinhas
Nuno
N
da Costa
Milton S
MS
D039903:Actinobacteria / Q000201:enzymology / Q000235:genetics*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001482:Base Composition; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000235:genetics; D015870:Gene Expression; D005798:Genes, Bacterial; D005964:Glucosyltransferases / Q000235:genetics / Q000378:metabolism*; D006156:Guanosine Diphosphate Sugars / Q000378:metabolism; D008969:Molecular Sequence Data; D010744:Phosphoric Monoester Hydrolases / Q000235:genetics / Q000378:metabolism*; D010802:Phylogeny; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D020539:Sequence Analysis, Protein; D013379:Substrate Specificity; D013403:Sugar Phosphates / Q000096:biosynthesis*; D014199:Trehalose / Q000031:analogs & derivatives* / Q000096:biosynthesis
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D006156:Guanosine Diphosphate Sugars; D011994:Recombinant Proteins; D013403:Sugar Phosphates; C082722:trehalose-6-phosphate; C100085:guanosine diphosphate glucose; D014199:Trehalose; D005964:Glucosyltransferases; C099692:trehalose synthase; C023180:trehalose-6-phosphate synthase; C023178:trehalose-phosphatase; D010744:Phosphoric Monoester Hydrolases