Rhizobium bacteria synthesize N-acylated In the symbiotic relationship between Rhizobium bacteria and leguminous host plants, a set of molecular signals is exchanged (1-3). The rhizobia have a set ofnodgenes that are required for the establishment of the root nodule symbiosis. These genes are grouped into three categories: common nod genes, required for nodulation and conserved in all of the nodulating bacteria; host-specific genes, which vary according to the bacterial cross-inoculation group; and regulatory nod genes, which activate nod gene expression, often in conjunction with plant-derived flavonoid inducers. The common and host-specific nod genes specify the production of signal molecules, the lipooligosaccharide Nod factors. The common nod genes are necessary for Nod factor production, while the host-specific nod genes are required for structural modifications which impart specificity to the various factors.The Nod factors ofdiverse rhizobia share certain structural features (2, 4-8). All are (3-1,4-N-acetyl-D-glucosamine (GlcNAc) oligosaccharides with various N-linked fatty acyl groups replacing the acetyl group on the nonreducing sugar residue. Rhizobium meliloti Nod factors also carry a 6-0 sulfate moiety on the reducing terminus (4) (Fig. 1). The presence of the sulfate group requires the nodH gene in R. melioti (9) and is a strong determinant ofthe specificity ofthe R. meliloti factors (9, 10). The nodH gene encodes an 0-sulfotransferase which can specifically transfer a sulfate to GlcN-,81,4-(GlcNAc)3 was prepared enzymatically with bovine P-1,4-galactosyltransferase from UDP-glucosamine and chitotriose. the reducing-end GlcNAc of a GlcNAc oligosaccharide (ref. 9 and unpublished data).The common nod genes were the first to be identified, because they are absolutely required for nodulation. Their central role in Nod factor production and in nodulation indicates that they encode unique and important functions. This study addresses the function of nodA and nodE, the first two genes in the common nod operon. It has been reported that nodB encodes an N-deacetylase that removes the nonreducing-end acetyl from a GlcNAc oligosaccharide (11). Here we report data that support this conclusion, and we show that nodA and nodB are required for an N-acyltransferase activity, which we detected using oligosaccharide substrates labeled with 35S by NodH-mediated sulfation. This activity would account for the acylation seen on the various rhizobial Nod factors. The activity is sensitive to oligosaccharide chain length, preferentially acting upon chitotetraose. To separate the activities of the deacetylase and acyltransferase, we synthesized an N-deacetylated substrate, 3. Use of this substrate obviates the need for nodB Abbreviation: ACP, acyl carrier protein.
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