Structural and enzymatic characterisation of the Type III effector NopAA (=GunA) from Sinorhizobium fredii USDA257 reveals a Xyloglucan hydrolase activity
Résumé
Rhizobia are nitrogen-fixing soil bacteria that can infect legume plants to establish root nodules symbiosis. To do that, a complex exchange of molecular signals occurs between plants and bacteria. Among them, rhizobial Nops (Nodulation outer proteins), secreted by a type III secretion system (T3SS) determine the host-specificity for efficient symbiosis with plant roots. Little is known about the molecular function of secreted Nops (also called effectors (T3E)) and their role in the symbiosis process. We performed the structure-function characterization of NopAA, a T3E from Sinorhizobium fredii by using a combination of X-ray crystallography, biochemical and biophysical approaches. this work displays for the first time a complete structural and biochemical characterization of a symbiotic T3E. Our results showed that NopAA has a catalytic domain with xyloglucanase activity extended by a N-terminal unfolded secretion domain that allows its secretion. We proposed that these original structural properties combined with the specificity of NopAA toward xyloglucan, a key component of root cell wall which is also secreted by roots in the soil, can give NopAA a strategic position to participate in recognition between bacteria and plant roots and to intervene in nodulation process. Many bacterial pathogens use type III secretion systems (T3SS) to inject virulence factors, named effectors, directly into the cytoplasm of target eukaryotic cells. Most of the T3SS apparatus components are conserved among plant and animal pathogens, suggesting a common mechanism of recognition and secretion of effec-tors (for review see 1). Secretion of effectors depends on the presence of secretion signals composed by 20 to 30 non-conserved amino acids at the N-terminus. Most of T3SS effectors (T3E) are predicted to possess a N-terminal intrinsically disordered (ID) region enriched in serine residues and required for secretion 2. Many secreted proteins also depend on the interaction with a cytoplasmic specific T3S chaperone 3 that stabilize ID regions in bacterial cytoplasm prior translocation. For SopB, a T3E of Salmonella it has been proposed that its cognate chaperone SigE may be responsible of the formation of ring like hexamers, conformation that may be required for substrate recognition by the type three apparatus 4. In pathogenic bacteria, T3SS genes coding for the secretion apparatus, effectors and accessory proteins are clustered in pathogenicity islands (PAIs) and organized in operons. Interestingly, this secretion strategy is also used by non-pathogenic organisms contributing to symbiotic interactions with hosts as shown for rhizobia 5,6. Rhizobia are soil bacteria having the ability to establish a specific symbiotic association with leguminous host-plant roots. This interaction leads to the formation of root nodules by the plant, specialized organs in which bacteria differentiate into nitrogen fixing bacteroids able to convert atmospheric nitrogen into ammonia for the usage in plant growth 7. This process starts with the secretion of
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