FtsZ condensates: An in vitro electron microscopy study

Abstract

In vivo cell division protein FtsZ from E. coli forms rings and spirals which have only been observed by low resolution light microscopy. We show that these suprastructures are likely formed by molecular crowding which is a predominant factor in prokaryotic cells and enhances the weak lateral bonds between proto-filaments. Although FtsZ assembles into single proto-filaments in dilute aqueous buffer, with crowding agents above a critical concentration, it forms polymorphic supramolecular structures including rings and toroids (with multiple protofilaments) about 200 nm in diameter, similar in appearance to DNA toroids, and helices with pitches of several hundred nm as well as long, linear bundles. Helices resemble those observed in vivo, whereas the rings and toroids may represent a novel energy minimized state of FtsZ, at a later stage of Z-ring constriction. We shed light on the molecular arrangement of FtsZ filaments within these suprastructures using high resolution electron microscopy.

Extreme C Terminus of Bacterial Cytoskeletal Protein FtsZ Plays Fundamental Role in Assembly Independent of Modulatory Proteins

“…3C). Such structures have previously been observed for Ec FtsZ only in the presence of higher concentrations of divalent cations or molecular crowding agents (7). The diameter of the Ec FtsZ CTVB rings was on average 370.64 Ϯ 46.24 nm with a width of 58.27 Ϯ 20.37 nm, somewhat larger than those observed for a similar structure composed of wild-type Bs FtsZ (183.41 Ϯ 34.27-nm diameter; 30.66 Ϯ 10.85-nm width) (supplemental Fig.…”

“…In vitro, FtsZ assembles into single-stranded polymers, or protofilaments, in the presence of GTP (4). Increasing the concentrations of divalent cations like Mg 2ϩ or Ca 2ϩ or adding crowding agents, such as methyl cellulose or DEAE-dextran to purified Escherichia coli and Bacillus subtilis FtsZ stimulates lateral interactions between protofilaments, leading to the formation of bundles, spirals, tubes, and sheets (5)(6)(7)(8).…”

In the Beginning, Escherichia coli Assembled the Proto-ring: An Initial Phase of Division

“…Such filaments have been observed when FtsZ is adsorbed onto mica surfaces, when calcium is present, or at low pH (Ͻ6.0) (29 -31). Long FtsZ filaments are also formed under more physiological conditions in the presence of accessory proteins or crowding agents that mimic the in vivo conditions (32)(33)(34). In solution, long FtsZ protofilaments have also been found assembled as ribbons on the outer surface of artificial membrane tubules (35).…”

Evidence That Bacteriophage λ Kil Peptide Inhibits Bacterial Cell Division by Disrupting FtsZ Protofilaments and Sequestering Protein Subunits

“…Hence, these basic structural units must further assemble into higher order structures to form the observed Z-ring in vivo (10,11). Indeed, protofilaments can be arranged in vitro into several higher order structures involving lateral interactions in the presence of crowding agents (7,12), calcium ions (13), or certain buffer conditions (14).…”