Demonstration by Membrane Reconstitution of a Butanol‐Soluble Intermediate in the Biosynthesis of the O9 Antigen of Escherichia coli

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Abstract
The activity in vitro of the mannan-synthesizing system of E. coli O9 depends on the presence of glucose in the growth medium of the bacteria. Inactive membranes of E. coli strain E988 grown without glucose gain mannan-synthesizing activity by reconstitution with a butanol extract obtained from the same bacteria grown with glucose. Inactive membranes could also be restored to biosynthetic activity by incubation with UDP-glucose in the presence of magnesium chloride. In this Mg-ion-dependent reaction, a glucolipid was formed which was extractable with butanol. It could be used for the reconstitution of inactive membranes. The products of incubations with GDP-mannose of reconstituted and active membranes were analyzed for electrophoretic mobility in sodium dodecylsulfate/polyacrylamide gel electrophoresis, molecular weight and composition. In all cases they proved to be the mannan attached to a hydrophobic mannose carrier, presumably a glucolipid. A glucolipid may be the intermediary mannose acceptor in the biosynthesis of the O9 antigen.