Characterization of the biosynthetic pathway of glucosylglycerate in the archaeon Methanococcoides burtonii.
The pathway for the synthesis of the organic solute glucosylglycerate (GG) is proposed based on the activities of the recombinant glucosyl-3-phosphoglycerate synthase (GpgS) and glucosyl-3-phosphoglycerate phosphatase (GpgP) from Methanococcoides burtonii. A mannosyl-3-phosphoglycerate phosphatase gene homologue (mpgP) was found in the genome of M. burtonii (http://www.jgi.doe.gov), but an mpgS gene coding for mannosyl-3-phosphoglycerate synthase (MpgS) was absent. The gene upstream of the mpgP homologue encoded a putative glucosyltransferase that was expressed in Escherichia coli. The recombinant product had GpgS activity, catalyzing the synthesis of glucosyl-3-phosphoglycerate (GPG) from GDP-glucose and d-3-phosphoglycerate, with a high substrate specificity. The recombinant MpgP protein dephosphorylated GPG to GG and was also able to dephosphorylate mannosyl-3-phosphoglycerate (MPG) but no other substrate tested. Similar flexibilities in substrate specificity were confirmed in vitro for the MpgPs from Thermus thermophilus, Pyrococcus horikoshii, and "Dehalococcoides ethenogenes." GpgS had maximal activity at 50 degrees C. The maximal activity of GpgP was at 50 degrees C with GPG as the substrate and at 60 degrees C with MPG. Despite the similarity of the sugar donors GDP-glucose and GDP-mannose, the enzymes for the synthesis of GPG or MPG share no amino acid sequence identity, save for short motifs. However, the hydrolysis of GPG and MPG is carried out by phosphatases encoded by homologous genes and capable of using both substrates. To our knowledge, this is the first report of the elucidation of a biosynthetic pathway for glucosylglycerate.
Costa
Joana
J
Departamento de Bioquímica and Centro de Neurociências e Biologia Celular, Universidade de Coimbra, 3001-401 Coimbra, Portugal. milton@ci.uc.pt.
Empadinhas
Nuno
N
Gonçalves
Luís
L
Lamosa
Pedro
P
Santos
Helena
H
da Costa
Milton S
MS
D001105:Archaea; D005947:Glucose / Q000031:analogs & derivatives / Q000096:biosynthesis; D005960:Glucosides / Q000096:biosynthesis*; D005964:Glucosyltransferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005988:Glyceric Acids / Q000378:metabolism; D017019:Methanosarcinaceae / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity
C509251:2-glucosylglycerate; D005960:Glucosides; D005988:Glyceric Acids; D011994:Recombinant Proteins; D005964:Glucosyltransferases; D017853:Phosphotransferases (Alcohol Group Acceptor); D005947:Glucose