Nano-encapsulated Escherichia coli Divisome Anchor ZipA, and in Complex with FtsZ

Abstract

The E. coli membrane protein ZipA, binds to the tubulin homologue FtsZ, in the early stage of cell division. We isolated ZipA in a Styrene Maleic Acid lipid particle (SMALP) preserving its position and integrity with native E. coli membrane lipids. Direct binding of ZipA to FtsZ is demonstrated, including FtsZ fibre bundles decorated with ZipA. Using Cryo-Electron Microscopy, small-angle X-ray and neutron scattering, we determine the encapsulated-ZipA structure in isolation, and in complex with FtsZ to a resolution of 1.6 nm. Three regions can be identified from the structure which correspond to, SMALP encapsulated membrane and ZipA transmembrane helix, a separate short compact tether, and ZipA globular head which binds FtsZ. The complex extends 12 nm from the membrane in a compact structure, supported by mesoscale modelling techniques, measuring the movement and stiffness of the regions within ZipA provides molecular scale analysis and visualisation of the early divisome.

ZipA Uses a Two-Pronged FtsZ-Binding Mechanism Necessary for Cell Division

“…10B ). This model would be consistent with the cryo-EM structure of full-length ZipA and FtsZ that places the globular domains of both proteins in close contact ( 20 ). Such a two-pronged binding model might also help explain why FtsA, which seemingly interacts only with the FtsZ CTP, has a lower binding affinity for FtsZ than ZipA.…”

“…Although a second study showed that oligomerization of FtsZ plays a significant role in strengthening ZipA’s apparent interaction with full-length FtsZ, the possibility of a secondary binding site could not be ruled out ( 8 ). Moreover, a recent cryo-EM structure of full-length ZipA in complex with FtsZ in lipid nanodiscs ( 20 ) showed that the globular domains of both proteins were closely associated, hinting at additional interactions beyond the canonical FtsZ CTP-binding site.…”

A comparison of SMA (styrene maleic acid) and DIBMA (di-isobutylene maleic acid) for membrane protein purification

“…In addition, SMALPs once formed are stable, and it is not necessary to supplement all buffers as with conventional detergents, making them technically easier to work with. SMALP purified proteins have successfully been used for both functional [9,12,13] and structural [16][17][18][19] studies. However SMA polymers do not solve all challenges associated with membrane proteins.…”

Exploring the dynamics of flagellar dynein within the axoneme with Fluctuating Finite Element Analysis

“…FFEA (Solernou et al ., 2018) represents globular macromolecules such as proteins as continuum objects (Oliver et al ., 2014), and has been previously used to model rotary ATPases, the ZipA/FtsZ membrane protein complexes, and cytoplasmic dynein (Richardson et al ., 2014; Lee et al ., 2019; Hanson et al ., 2020). Within the FFEA framework, proteins are approximated as viscoelastic solids (Wang and Zocchi, 2011) which change shape and explore conformational space under the effect of thermal fluctuations.…”