Mapping of the protein-coding regions of Rhizobium meliloti common nodulation genes

Abstract

An 8.5-kb EcoRI fragment containing the common nod region of the megaplasmid pRme41b of Rhizobium meliloti was recloned in plasmids of Escherichia coli, and a detailed restriction map was established. The region can express at least eight proteins in E. coli minicells and in an in vitro transcription/translation system, prepared from E. coli. Protein coding regions were determined by subcloning of restriction fragments, deletion mutations and by transposon mutagenesis. The coding regions for at least three polypeptide chains (mol. wts. 23 000, 28 500 and 44 000) were mapped on a 3.3-kb nod gene cluster. The 44 000 mol. wt. protein is expressed from a nod region, which is highly conserved in two Rhizobium species. The protein map of the 8.5-kb fragment was correlated to a map of insertion mutations with Nod and Fix phenotypes. The data suggest that the proteins encoded by the nod gene cluster may be involved in early steps of the nodulation process. Nod Fix symbiotic mutations were localized in the coding region for a 33 000 mol. wt. protein, suggesting that this polypeptide might be a fix gene product.