Isolation and characterization of Rhizobium tropici Nod factor sulfation genes

Rhizobium tropici produces a mixture of sulfated and non-sulfated Nod factors. The genes responsible for the sulfation process in R. tropici strain CFN299 were cloned and sequenced. These genes are homologous to the nodP, nodQ, and nodH genes from R. meliloti. The identity among the two species is 75% for nodP, 74% for nodQ, and 69% for nodH. NodH resembles sulfotransferases in general and NodQ has the characteristic purine-binding motifs and the PAPS 3'-phosphoadenosine 5'-phosphosulfate) motif. Mutants of NodP and NodH were obtained by site-directed mutagenesis. They are no longer able to synthesize the sulfated Nod factor, as was demonstrated in high-pressure liquid chromatography and thin-layer chromatography assays. The NodP- mutant had a decreased nodulation capacity in Phaseolus vulgaris Negro Xamapa bean plants. In contrast, NodH- and NodP- mutants acquired an increased capacity to nodulate the high-nitrogen-fixing bean cultivars N-8-116 and BAT-477. Nodulation was restored to normal levels when the mutants were complemented with a 16-kb clone carrying the wild-type genes. The role of the sulfate on Nod factors in R. tropici was dependent on the bean cultivar and the conditions assayed.