UDP-galactofuranose precursor required for formation of the lipopolysaccharide O antigen of Klebsiella pneumoniae serotype O1 is synthesized by the product of the rfbDKPO1 gene

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  1. Get@NRC: UDP-galactofuranose precursor required for formation of the lipopolysaccharide O antigen of Klebsiella pneumoniae serotype O1 is synthesized by the product of the rfbDKPO1 gene (Opens in a new window)
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TypeArticle
Journal titleJournal Of Biological Chemistry
Volume272
Issue7
Pages41214128; # of pages: 8
SubjectACID; amino acid; Amino Acid Sequence; AMINO-ACID; AMINO-ACID-SEQUENCE; analogs & derivatives; ANTIGEN; Antigens; backbone; bacterial; biosynthesis; Canada; carbohydrate; Carbohydrate Epimerases; chemistry; Cloning,Molecular; CLUSTER; DISEASE; Electrophoresis; ENCODES; ENZYME; Escherichia; Escherichia coli; ESCHERICHIA-COLI; Filtration; Form; FORMS; Galactose; GENE; Genes; genetics; immunology; Klebsiella; Klebsiella pneumoniae; LIPOPOLYSACCHARIDE; Lipopolysaccharide O Antigen; Magnetic Resonance Spectroscopy; metabolism; MOLECULAR; Molecular Sequence Data; Nadp; NMR; NMR spectroscopy; NMR-spectroscopy; O antigen; O Antigens; O-antigen; O1; PNEUMONIAE; POLYSACCHARIDE; protein; RESIDUES; SEQUENCE; Sequence Alignment; SEROTYPE; Serotyping; SPECTROSCOPY; structure; Support,Non-U.S.Gov't; Synthesis; UNIT; Uridine Diphosphate
AbstractThe O-side-chain polysaccharide in the lipopolysaccharide of Klebsiella pneumoniae O1 is based on a backbone structure of repeat units of [-->3)-beta-D-Galf-(1-->3)-alpha-D-Galp-(1-->]; this structure is termed D-galactan I. The rfb (O-antigen biosynthesis) gene cluster directs the synthesis of D-galactan I and consists of six genes termed rfbA-FKPO1. In this paper we show that rfbDKPO1 encodes a UDP-galactopyranose mutase (NAD(P)H-requiring) (EC 5.4.99. 9), which forms uridine 5'-(trihydrogen diphosphate) P'-alpha-D-galactofuranosyl ester (UDP-Galf), the biosynthetic precursor of galactofuranosyl residues. The deduced amino acid sequence of rfbDKPO1 shows 85% and 37.5% identity to the rfbDKPO8 gene of K. pneumoniae serotype O8 and the glf gene of Escherichia coli, respectively. The molecular mass of the purified RfbDKPO1 enzyme is 45 kDa as determined by SDS-polyacrylamide gel electrophoresis, while gel filtration revealed a molecular mass of 92 kDa, suggesting a dimeric structure for the native protein. The rfbDKPO1 gene product interconverts uridine 5'-(trihydrogen diphosphate) P'-alpha-D-galactopyranosyl ester (UDP-Galp) and UDP-Galf. Unlike Glf, RfbDKPO1 showed a requirement for NADH or NADPH, which could not be replaced by NAD or NADP. RfbDKPO1 was used to synthesize milligram quantities of UDP-Galf, allowing this compound to be purified and fully characterized in an intact form for the first time. The structure of UDP-Galf was proven by NMR spectroscopy
Publication date
LanguageEnglish
AffiliationNational Research Council Canada
Peer reviewedNo
NRC numberKOPLIN1997
NPARC number9387238
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Record identifier2da45715-4ae2-4bef-a1b5-7703d2d782f2
Record created2009-07-10
Record modified2016-05-09
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