Homo- and heterotypic interactions between Pss proteins involved in the exopolysaccharide transport system in Rhizobium leguminosarum bv. trifolii

Abstract

Rhizobium leguminosarum produces large amounts of exopolysaccharide (EPS) that has been shown to be an important determinant of successful nitrogen-fixing symbiosis with legume plants. EPS is assembled in a Wzx/Wzy-dependent manner, and proteins involved in the process are proposed to form a complex that enables coupling the synthesis of EPS subunits with their polymerization and transport. Pss proteins, which are encoded within the chromosomal polysaccharide synthesis cluster of Rhizobium leguminosarum bv. trifolii TA1, were subjected to interaction analysis. PssN was shown to form multimeric complexes in the outer membrane and interact with the extracellular PssO protein and the inner membrane oligomeric PssP co-polymerase. PssO was demonstrated to form oligomers in the presence of the cross-linker. Bacterial two-hybrid analysis showed that PssP interacts with PssL and PssT, counterparts of Gram-negative bacteria Wzx and Wzy proteins. Membrane topology of PssT is discussed in the context of its plausible Wzy-like polymerase activity, interactions with PssP and a possible impact of these interactions on EPS polymerization and chain length determination. The importance of protein-protein and putative protein-polysaccharide interactions in EPS transport is discussed. A topology model for the EPS transport system, with highlights on localization, functions and interactions between the Pss proteins, is proposed.