Sinorhizobium meliloti Sulfotransferase That Modifies Lipopolysaccharide

Abstract

ABSTRACT Sinorhizobium meliloti is a gram-negative soil bacterium found either in free-living form or as a nitrogen-fixing endosymbiont of a plant structure called the nodule. Symbiosis between S. meliloti and its plant host alfalfa is dependent on bacterial transcription of nod genes, which encode the enzymes responsible for synthesis of Nod factor. S. meliloti Nod factor is a lipochitooligosaccharide that undergoes a sulfate modification essential for its biological activity. Sulfate also modifies the carbohydrate substituents of the bacterial cell surface, including lipopolysaccharide (LPS) and capsular polysaccharide (K-antigen) (R. A. Cedergren, J. Lee, K. L. Ross, and R. I. Hollingsworth, Biochemistry 34: 4467-4477, 1995). We utilized the genomic sequence of S. meliloti to identify an open reading frame, SMc04267 (which we now propose to name lpsS ), which encodes an LPS sulfotransferase activity. We expressed LpsS in Escherichia coli and demonstrated that the purified protein functions as an LPS sulfotransferase. Mutants lacking LpsS displayed an 89% reduction in LPS sulfotransferase activity in vitro. However, lpsS mutants retain approximately wild-type levels of sulfated LPS when assayed in vivo, indicating the presence of an additional LPS sulfotransferase activity(ies) in S. meliloti that can compensate for the loss of LpsS. The lpsS mutant did show reduced LPS sulfation, compared to that of the wild type, under conditions that promote nod gene expression, and it elicited a greater number of nodules than did the wild type during symbiosis with alfalfa. These results suggest that sulfation of cell surface polysaccharides and Nod factor may compete for a limiting pool of intracellular sulfate and that LpsS is required for optimal LPS sulfation under these conditions.