Lipopolysaccharide biosynthesis in Rhizobium leguminosarum. Novel enzymes that process precursors containing 3-deoxy-D-manno-octulosonic acid.
The lipopolysaccharide of Rhizobium leguminosarum differs from that of other Gram-negative organisms. R. leguminosarum lipid A lacks phosphate groups, but it contains a galacturonic acid residue at the 4'-position and an aminogluconate moiety in place of the usual glucosamine 1-phosphate unit. R. leguminosarum lipid A is esterified with a peculiar long chain fatty acid, 27-hydroxyoctacosanoate, not found in enteric Gram-negative bacteria, and the inner core of R. leguminosarum contains mannose and galactose in place of heptose. Despite these differences, the biosynthesis of R. leguminosarum lipid A is initiated by the same seven enzyme pathway as in Escherichia coli (Raetz, C. R. H. (1993) J. Bacteriol. 175, 5745-5753) to form the phosphorylated precursor, (Kdo)2-lipid IVA, which is then processed differently. We now describe several novel Rhizobium-specific enzymes that recognize and modify (Kdo)2-lipid IVA. The 1- and 4'-phosphatases were detected using (Kdo)2-[1-32P]-lipid IVA and (Kdo)2-[4'-32P]-lipid IVA, respectively, as shown by release of 32Pi. In the presence of GDP-mannose and/or UDP-galactose, membranes of R. leguminosarum first transferred mannose and then galactose to (Kdo)2-[4'-32P]-lipid IVA. In addition, at least two hydrophobic metabolites were generated from (Kdo)2-[4'-32P]-lipid IVA in a manner that was dependent upon both membranes and a cytosolic factor from R. leguminosarum. These compounds are attributed to novel acylations of (Kdo)2-[4'-32P]-lipid IVA. E. coli membranes and cytosol did not catalyze any of the unique reactions detected in R. leguminosarum extracts. Our findings establish the conservation and versatility of (Kdo)2-lipid IVA as a lipid A precursor in bacteria.
Brozek
K A
KA
Department of Biochemistry, Duke University Medical Center, Durham, North Carolina 27710, USA. raetz@bchm.biochem.duke.edu
Kadrmas
J L
JL
Raetz
C R
CR
D000217:Acyltransferases / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism; D002240:Carbohydrate Sequence; D003600:Cytosol / Q000378:metabolism; D004926:Escherichia coli; D005700:Galactosyltransferases / Q000378:metabolism*; D006017:Glycolipids / Q000378:metabolism; D006155:Guanosine Diphosphate Mannose / Q000378:metabolism; D008050:Lipid A / Q000031:analogs & derivatives / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000096:biosynthesis*; D008364:Mannosyltransferases / Q000378:metabolism*; D008954:Models, Biological; D008969:Molecular Sequence Data; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D016961:Rhizobium leguminosarum / Q000201:enzymology*; D013400:Sugar Acids / Q000378:metabolism*; D014531:Uridine Diphosphate Galactose / Q000378:metabolism
D006017:Glycolipids; D008050:Lipid A; D008070:Lipopolysaccharides; D013400:Sugar Acids; C040142:lipid A precursors, bacterial; C002532:2-keto-3-deoxyoctonate; D014531:Uridine Diphosphate Galactose; D006155:Guanosine Diphosphate Mannose; D000255:Adenosine Triphosphate; D000217:Acyltransferases; D005700:Galactosyltransferases; D008364:Mannosyltransferases; D010749:Phosphoprotein Phosphatases