Nodulation protein NodL of Rhizobium leguminosarum O-acetylates lipo-oligosaccharides, chitin fragments and N-acetylglucosamine in vitro.
Upon induction of their nodulation genes, the root nodule-inducing Rhizobium bacteria produce lipo-oligosaccharide signal molecules. All lipo-oligosaccharides identified from Rhizobium leguminosarum bv. viciae carry an O-acetyl group at the C-6 position of the non-reducing terminal sugar, the presence of which is important for biological activity and host specificity. Previously we showed that a functional nodL gene product is required for the presence of this O-acetyl moiety. The production of polyclonal antibodies against isolated NodL protein, using a NodL-overproducing Escherichia coli strain is described. These antibodies were used (i) to elucidate the subcellular localization of the NodL protein, which appeared to be present in the cytosol, and (ii) for the purification of native NodL protein from E. coli. Here we provide biochemical proof that purified NodL protein has transacetylating activity in vitro with acetyl-CoA as the acetyl donor. NodL protein appeared to be able to acetylate various substrates, such as lipo-oligosaccharides, chitin fragments and N-acetylglucosamine. For chitinpentaose as the substrate we have shown, using mass spectrometry and NMR spectroscopy, that NodL protein substitutes one O-acetyl group at the C-6 position of the non-reducing terminal sugar.
Bloemberg
G V
GV
Institute of Molecular Plant Sciences, Leiden University, The Netherlands.
Thomas-Oates
J E
JE
Lugtenberg
B J
BJ
Spaink
H P
HP
D000107:Acetylation; D000117:Acetylglucosamine / Q000378:metabolism*; D000123:Acetyltransferases / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D000818:Animals; D001426:Bacterial Proteins / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D002686:Chitin / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000378:metabolism*; D011817:Rabbits; D016961:Rhizobium leguminosarum / Q000201:enzymology*; D013347:Subcellular Fractions / Q000201:enzymology
D001426:Bacterial Proteins; D008070:Lipopolysaccharides; D002686:Chitin; D000123:Acetyltransferases; D000117:Acetylglucosamine