The iaaL gene in the Pseudomonas syringae complex: functional characterization and biological activity

Abstract

Phytopathogenic bacteria of the Pseudomonas syringae complex are causal agents of diseases in a wide variety of woody and herbaceous plants with agronomic and ornamental interest. Indole-3-acetic acid (IAA) is an auxin phytohormone whose production is widely distributed among plant-associated bacteria. Some P. syringae strains can further metabolize IAA to the amino acid conjugate 3-indole-acetyl-ε-L-lysine (IAA-Lys), a process involving the enzyme IAA-Lys synthase, encoded by the iaaL gene. IAA-Lys is less biologically active than IAA, so it has been speculated that the conjugation of IAA with L-Lysine could allow the bacteria to control the levels of free IAA accumulated in the bacterial cytoplasm and/or secreted to the plant tissues. The iaaL gene is widespread in the P. syringae complex, and three different alleles (iaaLPsv, iaaLPsn and iaaLPto) have been described. Recently, we have identified a fourth allele (iaaLPsf) specifically encoded in the genome of strains isolated from Fraxinus excelsior. However, comparative analyses of the biochemical and biological activities of the different iaaL alleles have not been performed. In this work, the genomic context of these four alleles in a collection of P. syringae complex strains has been analyzed. In addition, we have constructed strains overexpressing each of these iaaL alleles and analysed their biological activities using an elongation assay of Arabidopsis thaliana roots. Finally, expression of these alleles in E. coli allowed the purification of these four IaaL proteins and the analysis of their specific activities using an in vitro enzymatic coupling assay.