Protein interactions involved in cyclic beta-1,2-glucans metabolism

L. SOLEDAD GUIDOLIN; SUSANA MORRONE; FRANCISCO F. GUAIMAS; DIEGO J. COMERCI; ANDRÉS E. CIOCCHINI

Ciudad de Buenos Aires

Simposio; GlycoAR 2014; 2014

Cyclic beta-1,2-glucans (CBG) are periplasmic homopolysaccharides that play an important role in several symbiotic and pathogenic relationships. Brucella abortus CBG synthase (Cgs) is an integral inner membrane (IIM) protein which catalyzes the synthesis of CBG. Once synthesized in the cytoplasm, CBG are transported to the periplasm by the CBG transporter (Cgt) and succinylated by the CBG modifier enzyme (Cgm). Cgt and Cgm, as well as Cgs, are IIM proteins. In this work, we used a bacterial two-hybrid system and co-immunoprecipitation techniques to study the interaction network between these three IIM proteins. Our results indicate that Cgs interacts with Cgt and Cgm, and that Cgt interacts with Cgm. We also observe that each one of these proteins form homotypic complexes. Analyses carried out with Cgs in-frame insertion mutants and a Cgs deletion mutant revealed that a coiled-coil motif located in the NH-terminal domain of the protein is important to sustain protein interactions, although other regions may also be implicated. Finally, we performed analyses of fluorescence microscopy and found that Cgs and Cgm are localized at the cell poles in B. abortus. We propose that Cgs, Cgt and Cgm form an inner membrane protein complex, probably located at the poles of bacteria, necessary to coordinate the synthesis, transport to periplasm and succinylation of CBG.