Prediction and functional analysis of GGDEF/EAL domain-containing proteins in Azorhizobium caulinodans ORS571

Abstract:

[Objective] c-di-GMP, an important second messenger regulating multiple functions of bacteria, is generally synthesized and hydrolysed by proteins containing GGDEF or EAL domain. In this study, we analyzed the genome-wide GGDEF/EAL domain-containing proteins of Azorhizobium caulinodans ORS571, and selected three GGDEF-EAL composite proteins (AZC_3085, AZC_3226 and AZC_4658) for functional analysis. [Methods] SMART and CLUSTALW were used for prediction and multi-alignment of GGDEF/EAL domain-containing proteins. Mutants were constructed by homologous recombination. Phenotypes including cell motility, exopolysaccharide (EPS) production, biofilm formation and nodulation with legume host were investigated. [Results] There were 37 GGDEF/EAL domain-containing proteins in A. caulinodans ORS571. Mutant △4658 showed deficiency in cell motility, while its EPS production and biofilm formation were higher than that of wild type. Mutant △4658 showed stronger competitiveness than wild type in competitive nodulation assay. The loss of AZC_4658 led to the increase of intracellular c-di-GMP level. Mutants △3085 and △3226 did not show obvious difference in comparison with wild type. [Conclusion] The vast number of GGDEF/EAL domain-containing proteins suggested that c-di-GMP may play an important role in signal transduction of ORS571. The GGDEF-EAL composite protein AZC_4658 was involved in cell motility, EPS production, biofilm formation and nodulation of A. caulinodans ORS571.