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

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

Buenos Aires

Simposio; 1er Simposio Argentino de Glicobiología GlycoAR 2014; 2014

Cyclic -1,2-glucans (CG) are periplasmic homopolysaccharides that play an important role in several symbiotic and pathogenic relationships. Brucella abortus CG synthase (Cgs) is an integral inner membrane (IIM) protein which catalyzes the synthesis of CG. Once synthesized in the cytoplasm, CG are transported to the periplasm by the CβG transporter (Cgt) and succinylated by the CβG 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 CG.