Canavanine utilization via homoserine and hydroxyguanidine by a PLP-dependent γ-lyase in Pseudomonadaceae and Rhizobiales.

Canavanine, the δ-oxa-analogue of arginine, is produced as one of the main nitrogen storage compounds in legume seeds and has repellent properties. Its toxicity originates from incorporation into proteins as well as arginase-mediated hydrolysis to canaline that forms stable oximes with carbonyls. So far no pathway or enzyme has been identified acting specifically on canavanine. Here we report the characterization of a novel PLP-dependent enzyme, canavanine-γ-lyase, that catalyzes the elimination of hydroxyguanidine from canavanine to subsequently yield homoserine. Homoserine-dehydrogenase, aspartate-semialdehyde-dehydrogenase and ammonium-aspartate-lyase activities are also induced for facilitating canavanine utilization. We demonstrate that this novel pathway is found in certain Pseudomonas species and the Rhizobiales symbionts of legumes. The findings broaden the diverse reactions that the versatile class of PLP-dependent enzymes is able to catalyze. Since canavanine utilization is found prominently in root-associated bacteria, it could have important implications for the establishment and maintenance of the legume rhizosphere.

Pseudomonas canavaninivorans sp. nov., isolated from bean rhizosphere.

A novel canavanine-degrading bacterium, strain HB002T, was isolated from rhizosphere soil of a catch crop field collected from the island of Reichenau in Konstanz, Germany, and characterized by using polyphasic taxonomy. The facultative aerobe, rod-shaped, Gram-stain-negative bacterium was oxidase- and catalase-positive. The isolate was able to grow on canavanine as a sole carbon and nitrogen source. Results of phylogenetic analysis based on 16S rRNA gene sequences revealed highest similarities to Pseudomonas bijieensis (L22-9T, 99.93 %), Pseudomonas brassicacearum subsp. neoaurantiaca (ATCC 49054T, 99.76 %), Pseudomonas brassicacearum subsp. brassicacearum (DBK 11T, 99.63 %), Pseudomonas thivervalensis (DSM 13194T, 99.51 %), Pseudomonas kilonensis (DSM 13647T, 99.39 %) and Pseudomonas corrugata (ATCC29736T, 99.39 %). Marker gene analysis placed the strain in the intrageneric group of Pseudomonas fluorescens, subgroup P. corrugata. In silico DNA-DNA hybridization and average nucleotide identity values were both under the recommended thresholds for species delineation. The predominant fatty acids of strain HB002T were C16 : 0, C17 : 0 cyclo ω7c and C18 : 1 ω7c. The major respiratory quinone was Q9, followed by Q8 and minor components of Q7 and Q10. Results from the phenotypic characterization showd the strain's inability to hydrolyse gelatin and to assimilate N-acetyl glucosamide and a positive enzymatic activity of acid phosphatase and naphthol-AS-BI phosphohydrolase that distinguish this strain from closely related type strains. Taken together, these results show that strain HB002T represents a novel species in the genus Pseudomonas, for which the name Pseudomonas canavaninivorans sp. nov. is proposed. The type strain is HB002T (=DSM 112525T=LMG 32336T).