Structural Organization of FtsB, a Transmembrane Protein of the Bacterial Divisome

Abstract

We report the first structural analysis of an integral membrane protein of the bacterial divisome. FtsB is a single-pass membrane protein with a periplasmic coiled coil. Its heterologous association with its partner FtsL represents an essential event for the recruitment of the late components to the division site. Using a combination of mutagenesis, computational modeling and X-ray crystallography, we determined that FtsB self-associates and we investigated its structural organization. We found that the transmembrane domain of FtsB homo-oligomerizes through an evolutionarily conserved interaction interface where a polar residue (Gln 16) plays a critical role through the formation of an inter-helical hydrogen bond. The crystal structure of the periplasmic domain, solved as a fusion with Gp7, shows that 30 juxta-membrane amino acids of FtsB form a canonical coiled coil. The presence of conserved Gly residue in the linker region suggests that flexibility between the transmembrane and coiled coil domains is functionally important. We hypothesize that the transmembrane helices of FtsB form a stable dimeric core for its association with FtsL into a higher-order oligomer, and that FtsL is required to stabilize the periplasmic domain of FtsB, leading to the formation of a complex that is competent for binding to FtsQ, and to their consequent recruitment to the divisome. The study provides an experimentally validated structural model and identifies point mutations that disrupt association, thereby establishing important groundwork for the functional characterization of FtsB in vivo.

The Transmembrane Domains of the Bacterial Cell Division Proteins FtsB and FtsL Form a Stable High-Order Oligomer

“…The data shows that the TM region of the FtsB-FtsL complex is stably folded and that the TM helices are likely a major determinant for the association of the complex. We also show that FtsL and FtsB forms a 1:1 higher-order oligomeric complex, which is consistent with our previous hypothesis that the FtsB-FtsL complex is likely a tetramer 18 (Figure 1c). From these results, we hypothesize that the FtsB-FtsL complex may form a multi-valent binding hub that is important for the stabilization of the Z-ring.…”

“…Previous studies indicate that both the transmembrane and the coiled coil domains are involved in their mutual association 6,7,17 . To investigate this we recently studied the structural organization of the individual domains of FtsB in isolation 18 . Using an in vivo interaction assay, we showed that the TM domain of FtsB self-associates in E. coli membranes.…”

Fine-mapping the Contact Sites of the Escherichia coli Cell Division Proteins FtsB and FtsL on the FtsQ Protein*

“…The complex can be studied using minimally altered, functional, full-length proteins in their natural environment. Extending the current data with photo cross-linking from FtsB and FtsL could provide clues concerning the FtsQBL stoichiometry and test, for instance, the recently reported structural organization of FtsB in vivo within the context of the membrane-spanning complex (44). Other divisome proteins can be added to the system to probe for other interaction interfaces, for instance at the site of the conserved DLRY(d/e)(s/t)G motif.…”

The Soluble Periplasmic Domains of Escherichia coli Cell Division Proteins FtsQ/FtsB/FtsL Form a Trimeric Complex with Submicromolar Affinity

“…Recently, the crystal structure of a fragment of FtsB comprising 30 membrane-proximal periplasmic amino acids was solved as a fusion with Gp7 showing a homodimeric helical form (38). However, the thermal stability of this dimer was low due to the few hydrogen bonds present.…”

“…Probably, the TM segments of the full-length FtsB and FtsL proteins are required for efficient interaction. The TMs may extend the predicted coiled coil structures in the respective periplasmic domains that are located directly adjacent to the membrane (11,18,38). To compensate for the absence of potentially interacting TM segments in our constructs, we fused artificial coils of opposite charge to the N terminus, a strategy that has proven successful to dimerize the soluble domains of DivIC and FtsL from Streptococcus pneumoniae that show some similarity to FtsB and FtsL of E. coli, respectively (24,39).…”