BACTERIAL NODULATION PROTEIN NODZ IS A CHITIN OLIGOSACCHARIDE FUCOSYL-TRANSFERASE WHICH CAN ALSO RECOGNIZE RELATED SUBSTRATES OF ANIMAL ORIGIN

The nodZ gene, which is present in various soil bacteria such as Brady rhizobium japonicum, Azorhizobium caulinodans, and Rhizobium loti, is involved in the addition of a fucosyl residue to the reducing N-acetyl glucosamine residue of lipochitin oligosaccharide (LCO) signal molecul es, Using an Escherichia coli strain that produces large quantities of the NodZ protein of B. japonicum, we have purified the NodZ protein t o homogeneity The purified NodZ protein appears to be active in an in vitro transfucosylation assay in which GDP-beta-fucose and LCOs or chi tin oligosaccharides are used as substrates, The products of the in vi tro reaction using chitin oligosaccharides as substrate were studied b y using mass spectrometry, linkage analysis, and composition analysis, The data show that one fucose residue is added to C6 of the reducing- terminal N-acetylglucosamine residue, The substrate specificity of Nod Z protein was analyzed in further detail, using radiolabeled GDP-beta- fucose as the donor, The results show that chitin oligosaccharides are much better substrates than LCOs, suggesting that in Rhizobium NodZ f ucosylates chitin oligosaccharides prior to their acylation, The free glycan core pentasaccharides of N-linked glycoproteins are also substr ates for NodZ. Therefore, the NodZ enzymes seems to have an activity e quivalent to that of the enzyme involved in the addition of the Cb-lin ked fucosyl substituent in the glycan core of N-linked glycoproteins i n eukaryotes, Oligosaccharides that contain only one N-acetylglucosami ne at the reducing terminus are also substrates for NodZ, although in this case very high concentrations of such oligosaccharides are needed , An example is the leukocyte antigen Lewis-X, which can be converted by NodZ to a novel fucosylated derivative that could be used for bindi ng studies with E-selectin.