scholarly journals Structure and Mutational Analyses of Escherichia coli ZapD Reveal Charged Residues Involved in FtsZ Filament Bundling

2016 ◽  
Vol 198 (11) ◽  
pp. 1683-1693 ◽  
Author(s):  
Elyse J. Roach ◽  
Charles Wroblewski ◽  
Laura Seidel ◽  
Alison M. Berezuk ◽  
Dyanne Brewer ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Cell Division ◽  
Bacterial Cell ◽  
Site Directed Mutagenesis ◽  
Bacterial Cell Division ◽  
Wild Type ◽  
Content Type ◽  
E Coli ◽  
Z Ring

ABSTRACTBacterial cell division is an essential and highly coordinated process. It requires the polymerization of the tubulin homologue FtsZ to form a dynamic ring (Z-ring) at midcell. Z-ring formation relies on a group of FtsZ-associatedproteins (Zap) for stability throughout the process of division. InEscherichia coli, there are currently five Zap proteins (ZapA through ZapE), of which four (ZapA, ZapB, ZapC, and ZapD) are small soluble proteins that act to bind and bundle FtsZ filaments. In particular, ZapD forms a functional dimer and interacts with the C-terminal tail of FtsZ, but little is known about its structure and mechanism of action. Here, we present the crystal structure ofEscherichia coliZapD and show it forms a symmetrical dimer with centrally located α-helices flanked by β-sheet domains. Based on the structure of ZapD and its chemical cross-linking to FtsZ, we targeted nine charged ZapD residues for modification by site-directed mutagenesis. Usingin vitroFtsZ sedimentation assays, we show that residues R56, R221, and R225 are important for bundling FtsZ filaments, while transmission electron microscopy revealed that altering these residues results in different FtsZ bundle morphology compared to those of filaments bundled with wild-type ZapD. ZapD residue R116 also showed altered FtsZ bundle morphology but levels of FtsZ bundling similar to that of wild-type ZapD. Together, these results reveal that ZapD residues R116, R221, and R225 likely participate in forming a positively charged binding pocket that is critical for bundling FtsZ filaments.IMPORTANCEZ-ring assembly underpins the formation of the essential cell division complex known as the divisome and is required for recruitment of downstream cell division proteins. ZapD is one of several proteins inE. colithat associates with the Z-ring to promote FtsZ bundling and aids in the overall fitness of the division process. In the present study, we describe the dimeric structure ofE. coliZapD and identify residues that are critical for FtsZ bundling. Together, these results advance our understanding about the formation and dynamics of the Z-ring prior to bacterial cell division.

Localization, Assembly, and Activation of the Escherichia coli Cell Division Machinery

EcoSal Plus ◽  
2021 ◽  
Author(s):  
Petra Anne Levin ◽  
Anuradha Janakiraman
Keyword(s):  
Escherichia Coli ◽  
Cell Division ◽  
Bacterial Cell ◽  
Coli Cell ◽  
Bacterial Cell Division ◽  
Content Type ◽  
E Coli ◽  
Insight Into

Decades of research, much of it in Escherichia coli , have yielded a wealth of insight into bacterial cell division. Here, we provide an overview of the E. coli division machinery with an emphasis on recent findings.

scholarly journals DrpB (YedR) Is a Nonessential Cell Division Protein in Escherichia coli

2020 ◽  
Vol 202 (23) ◽  
Author(s):  
Atsushi Yahashiri ◽  
Jill T. Babor ◽  
Ariel L. Anwar ◽  
Ryan P. Bezy ◽  
Evan W. Piette ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Cell Division ◽  
Ionic Strength ◽  
Bacterial Cell ◽  
Enteric Bacteria ◽  
High Ionic Strength ◽  
Bacterial Cell Division ◽  
Content Type ◽  
E Coli ◽  
Low Ionic Strength

ABSTRACT We report that the small Escherichia coli membrane protein DrpB (formerly YedR) is involved in cell division. We discovered DrpB in a screen for multicopy suppressors of a ΔftsEX mutation that prevents divisome assembly when cells are plated on low ionic strength medium, such as lysogeny broth without NaCl. Characterization of DrpB revealed that (i) translation initiates at an ATG annotated as codon 22 rather than the GTG annotated as codon 1, (ii) DrpB localizes to the septal ring when cells are grown in medium of low ionic strength but localization is greatly reduced in medium of high ionic strength, (iii) overproduction of DrpB in a ΔftsEX mutant background improves recruitment of the septal peptidoglycan synthase FtsI, implying multicopy suppression works by rescuing septal ring assembly, (iv) a ΔdrpB mutant divides quite normally, but a ΔdrpB ΔdedD double mutant has a strong division and viability defect, albeit only in medium of high ionic strength, and (v) DrpB homologs are found in E. coli and a few closely related enteric bacteria, but not outside this group. In sum, DrpB is a poorly conserved nonessential division protein that improves the efficiency of cytokinesis under suboptimal conditions. Proteins like DrpB are likely to be a widespread feature of the bacterial cell division apparatus, but they are easily overlooked because mutants lack obvious shape defects. IMPORTANCE A thorough understanding of bacterial cell division requires identifying and characterizing all of the proteins that participate in this process. Our discovery of DrpB brings us one step closer to this goal in E. coli.

scholarly journals Of the Morphogenes that Make a Ring, A Rod and a Sphere in Escherichia Coli

2007 ◽  
Vol 90 (2-3) ◽  
pp. 59-72 ◽  
Author(s):  
Medhatm Khattar ◽  
Issmat I. Kassem ◽  
Ziad W. El-Hajj
Keyword(s):  
Escherichia Coli ◽  
Cell Division ◽  
Bacterial Cell ◽  
Bacterial Cell Division ◽  
Antibacterial Compounds ◽  
E Coli ◽  
The Past ◽  
New Knowledge ◽  
Fundamental Process ◽  
Simple Question

In 1993, William Donachie wrote “The success of molecular genetics in the study of bacterial cell division has been so great that we find ourselves, armed with much greater knowledge of detail, confronted once again with the same naive questions that we set to answer in the first place”1. Indeed, attempts to answer the apparently simple question of how a bacterial cell divides have led to a wealth of new knowledge, in particular over the past decade and a half. And while some questions have been answered to a great extent since the early reports of isolation of division mutants of Escherichia coli2,3, some key pieces of the puzzle remain elusive. In addition to it being a fundamental process in bacteria that merits investigation in its own right, studying the process of cell division offers an abundance of new targets for the development of new antibacterial compounds that act directly against key division proteins and other components of the cytoskeleton, which are encoded by the morphogenes of E. coli4. This review aims to present the reader with a snapshot summary of the key players in E. coli morphogenesis with emphasis on cell division and the rod to sphere transition.

scholarly journals High-resolution crystal structures of Escherichia coli FtsZ bound to GDP and GTP

2020 ◽  
Vol 76 (2) ◽  
pp. 94-102 ◽  
Author(s):  
Maria A. Schumacher ◽  
Tomoo Ohashi ◽  
Lauren Corbin ◽  
Harold P. Erickson
Keyword(s):  
Escherichia Coli ◽  
Staphylococcus Aureus ◽  
Cell Division ◽  
High Resolution ◽  
Crystal Structures ◽  
Bacterial Cell ◽  
Model System ◽  
E Coli ◽  
Z Ring ◽  
Main Model

Bacterial cytokinesis is mediated by the Z-ring, which is formed by the prokaryotic tubulin homolog FtsZ. Recent data indicate that the Z-ring is composed of small patches of FtsZ protofilaments that travel around the bacterial cell by treadmilling. Treadmilling involves a switch from a relaxed (R) state, favored for monomers, to a tense (T) conformation, which is favored upon association into filaments. The R conformation has been observed in numerous monomeric FtsZ crystal structures and the T conformation in Staphylococcus aureus FtsZ crystallized as assembled filaments. However, while Escherichia coli has served as a main model system for the study of the Z-ring and the associated divisome, a structure has not yet been reported for E. coli FtsZ. To address this gap, structures were determined of the E. coli FtsZ mutant FtsZ(L178E) with GDP and GTP bound to 1.35 and 1.40 Å resolution, respectively. The E. coli FtsZ(L178E) structures both crystallized as straight filaments with subunits in the R conformation. These high-resolution structures can be employed to facilitate experimental cell-division studies and their interpretation in E. coli.

scholarly journals Analysis of MinC Reveals Two Independent Domains Involved in Interaction with MinD and FtsZ

2000 ◽  
Vol 182 (14) ◽  
pp. 3965-3971 ◽  
Author(s):  
Zonglin Hu ◽  
Joe Lutkenhaus
Keyword(s):  
Escherichia Coli ◽  
Cell Division ◽  
Functional Domains ◽  
Wild Type ◽  
Terminal Domain ◽  
Ring Assembly ◽  
Z Ring ◽  
Ftsz Assembly

ABSTRACT In Escherichia coli FtsZ assembles into a Z ring at midcell while assembly at polar sites is prevented by themin system. MinC, a component of this system, is an inhibitor of FtsZ assembly that is positioned within the cell by interaction with MinDE. In this study we found that MinC consists of two functional domains connected by a short linker. When fused to MalE the N-terminal domain is able to inhibit cell division and prevent FtsZ assembly in vitro. The C-terminal domain interacts with MinD, and expression in wild-type cells as a MalE fusion disrupts minfunction, resulting in a minicell phenotype. We also find that MinC is an oligomer, probably a dimer. Although the C-terminal domain is clearly sufficient for oligomerization, the N-terminal domain also promotes oligomerization. These results demonstrate that MinC consists of two independently functioning domains: an N-terminal domain capable of inhibiting FtsZ assembly and a C-terminal domain responsible for localization of MinC through interaction with MinD. The fusion of these two independent domains is required to achieve topological regulation of Z ring assembly.

scholarly journals MinC, MinD, and MinE Drive Counter-oscillation of Early-Cell-Division Proteins Prior to Escherichia coli Septum Formation

mBio ◽  
2013 ◽  
Vol 4 (6) ◽  
Author(s):  
Paola Bisicchia ◽  
Senthil Arumugam ◽  
Petra Schwille ◽  
David Sherratt
Keyword(s):  
Escherichia Coli ◽  
Cell Division ◽  
Lipid Bilayers ◽  
Epifluorescence Microscopy ◽  
Bacterial Cell Division ◽  
Content Type ◽  
Septum Formation ◽  
Early Stages ◽  
Z Ring ◽  
High Concentrations

ABSTRACTBacterial cell division initiates with the formation of a ring-like structure at the cell center composed of the tubulin homolog FtsZ (the Z-ring), which acts as a scaffold for the assembly of the cell division complex, the divisome. Previous studies have suggested that the divisome is initially composed of FtsZ polymers stabilized by membrane anchors FtsA and ZipA, which then recruit the remaining division proteins. The MinCDE proteins prevent the formation of the Z-ring at poles by oscillating from pole to pole, thereby ensuring that the concentration of the Z-ring inhibitor, MinC, is lowest at the cell center. We show that prior to septum formation, the early-division proteins ZipA, ZapA, and ZapB, along with FtsZ, assemble into complexes that counter-oscillate with respect to MinC, and with the same period. We propose that FtsZ molecules distal from high concentrations of MinC form relatively slowly diffusing filaments that are bound by ZapAB and targeted to the inner membrane by ZipA or FtsA. These complexes may facilitate the early stages of divisome assembly at midcell. As MinC oscillates toward these complexes, FtsZ oligomerization and bundling are inhibited, leading to shorter or monomeric FtsZ complexes, which become less visible by epifluorescence microscopy because of their rapid diffusion. Reconstitution of FtsZ-Min waves on lipid bilayers shows that FtsZ bundles partition away from high concentrations of MinC and that ZapA appears to protect FtsZ from MinC by inhibiting FtsZ turnover.IMPORTANCEA big issue in biology for the past 100 years has been that of how a cell finds its middle. InEscherichia coli, over 20 proteins assemble at the cell center at the time of division. We show that the MinCDE proteins, which prevent the formation of septa at the cell pole by inhibiting FtsZ, drive the counter-oscillation of early-cell-division proteins ZapA, ZapB, and ZipA, along with FtsZ. We propose that FtsZ forms filaments at the pole where the MinC concentration is the lowest and acts as a scaffold for binding of ZapA, ZapB, and ZipA: such complexes are disassembled by MinC and reform within the MinC oscillation period before accumulating at the cell center at the time of division. The ability of FtsZ to be targeted to the cell center in the form of oligomers bound by ZipA and ZapAB may facilitate the early stages of divisome assembly.

scholarly journals Architecture of the ring formed by the tubulin homologue FtsZ in bacterial cell division

eLife ◽  
2014 ◽  
Vol 3 ◽  
Author(s):  
Piotr Szwedziak ◽  
Qing Wang ◽  
Tanmay A M Bharat ◽  
Matthew Tsim ◽  
Jan Löwe
Keyword(s):  
Cell Division ◽  
Molecular Model ◽  
Three Dimensional ◽  
Bacterial Cell Division ◽  
Electron Cryomicroscopy ◽  
E Coli ◽  
Ftsz Ring ◽  
Z Ring

Membrane constriction is a prerequisite for cell division. The most common membrane constriction system in prokaryotes is based on the tubulin homologue FtsZ, whose filaments in E. coli are anchored to the membrane by FtsA and enable the formation of the Z-ring and divisome. The precise architecture of the FtsZ ring has remained enigmatic. In this study, we report three-dimensional arrangements of FtsZ and FtsA filaments in C. crescentus and E. coli cells and inside constricting liposomes by means of electron cryomicroscopy and cryotomography. In vivo and in vitro, the Z-ring is composed of a small, single-layered band of filaments parallel to the membrane, creating a continuous ring through lateral filament contacts. Visualisation of the in vitro reconstituted constrictions as well as a complete tracing of the helical paths of the filaments with a molecular model favour a mechanism of FtsZ-based membrane constriction that is likely to be accompanied by filament sliding.

scholarly journals Cooperative ordering of treadmilling filaments in cytoskeletal networks of FtsZ and its crosslinker ZapA

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Paulo Caldas ◽  
Mar López-Pelegrín ◽  
Daniel J. G. Pearce ◽  
Nazmi Burak Budanur ◽  
Jan Brugués ◽  
...  
Keyword(s):  
Cell Division ◽  
Bacterial Cell ◽  
Large Scale ◽  
Bacterial Cell Division ◽  
Division Site ◽  
Associated Proteins ◽  
Z Ring ◽  
Cytoskeletal Networks ◽  
Cooperative Ordering

AbstractDuring bacterial cell division, the tubulin-homolog FtsZ forms a ring-like structure at the center of the cell. This Z-ring not only organizes the division machinery, but treadmilling of FtsZ filaments was also found to play a key role in distributing proteins at the division site. What regulates the architecture, dynamics and stability of the Z-ring is currently unknown, but FtsZ-associated proteins are known to play an important role. Here, using an in vitro reconstitution approach, we studied how the well-conserved protein ZapA affects FtsZ treadmilling and filament organization into large-scale patterns. Using high-resolution fluorescence microscopy and quantitative image analysis, we found that ZapA cooperatively increases the spatial order of the filament network, but binds only transiently to FtsZ filaments and has no effect on filament length and treadmilling velocity. Together, our data provides a model for how FtsZ-associated proteins can increase the precision and stability of the bacterial cell division machinery in a switch-like manner.

scholarly journals Antimicrobial Activity Evaluation of Tebipenem (SPR859), an Orally Available Carbapenem, against a Global Set of Enterobacteriaceae Isolates, Including a Challenge Set of Organisms

2019 ◽  
Vol 63 (6) ◽  
Author(s):  
S. J. Ryan Arends ◽  
Paul R. Rhomberg ◽  
Nicole Cotroneo ◽  
Aileen Rubio ◽  
Robert K. Flamm ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Antimicrobial Activity ◽  
Urinary Tract Infection ◽  
Urinary Tract ◽  
Tract Infection ◽  
Broth Microdilution ◽  
Wild Type ◽  
Content Type ◽  
E Coli

ABSTRACT The antimicrobial activity of tebipenem and other carbapenem agents were tested in vitro against a set of recent clinical isolates responsible for urinary tract infection (UTI), as well as against a challenge set. Isolates were tested by reference broth microdilution and included Escherichia coli (101 isolates), Klebsiella pneumoniae (208 isolates), and Proteus mirabilis (103 isolates) species. Within each species tested, tebipenem showed equivalent MIC50/90 values to those of meropenem (E. coli MIC50/90, ≤0.015/0.03 mg/liter; K. pneumoniae MIC50/90, 0.03/0.06 mg/liter; and P. mirabilis MIC50/90, 0.06/0.12 mg/liter) and consistently displayed MIC90 values 8-fold lower than imipenem. Tebipenem and meropenem (MIC50, 0.03 mg/liter) showed equivalent MIC50 results against wild-type, AmpC-, and/or extended-spectrum β-lactamase (ESBL)-producing isolates. Tebipenem also displayed MIC50/90 values 4- to 8-fold lower than imipenem against the challenge set. All carbapenem agents were less active (MIC50, ≥8 mg/liter) against isolates carrying carbapenemase genes. These data confirm the in vitro activity of the orally available agent tebipenem against prevalent UTI Enterobacteriaceae species, including those producing ESBLs and/or plasmid AmpC enzymes.

scholarly journals ISKpn40-Mediated Mobilization of the Colistin Resistance Gene mcr-3.11 in Escherichia coli

2020 ◽  
Vol 64 (10) ◽  
Author(s):  
Yu-Zhang He ◽  
Teng-Fei Long ◽  
Cai-Ping Chen ◽  
Bing He ◽  
Xing-Ping Li ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Resistance Gene ◽  
Wild Type ◽  
Colistin Resistance ◽  
Content Type ◽  
First Report ◽  
E Coli ◽  
Target Sites

ABSTRACT The mobile colistin resistance gene mcr-3 has globally disseminated since it was first reported in 2017 in Escherichia coli. In vitro mobilization assays in this study demonstrate the functionality of the composite transposon structure ISKpn40-mcr-3.11-dgkA-ISKpn40 in wild-type and recA− E. coli strains. These transpositions generated 4-bp duplications at the target sites. This is the first report demonstrating the mobility of the mcr-3.11 gene by transposition.

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