scholarly journals Overexpression of lpxT Gene in Escherichia coli Inhibits Cell Division and Causes Envelope Defects without Changing the Overall Phosphorylation Level of Lipid A

2020 ◽  
Vol 8 (6) ◽  
pp. 826
Author(s):  
Federica A. Falchi ◽  
Flaviana Di Lorenzo ◽  
Roberto Pizzoccheri ◽  
Gianluca Casino ◽  
Moira Paroni ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Cell Division ◽  
Lipid A ◽  
Cell Envelope ◽  
Osmotic Shock ◽  
Ectopic Expression ◽  
Phenotypic Effect ◽  
Osmotic Lysis ◽  
Membrane Defects ◽  
Inner Membrane Protein

LpxT is an inner membrane protein that transfers a phosphate group from the essential lipid undecaprenyl pyrophosphate (C-55PP) to the lipid A moiety of lipopolysaccharide, generating a lipid A tris-phosphorylated species. The protein is encoded by the non-essential lpxT gene, which is conserved in distantly related Gram-negative bacteria. In this work, we investigated the phenotypic effect of lpxT ectopic expression from a plasmid in Escherichia coli. We found that lpxT induction inhibited cell division and led to the formation of elongated cells, mostly with absent or altered septa. Moreover, the cells became sensitive to detergents and to hypo-osmotic shock, indicating that they had cell envelope defects. These effects were not due to lipid A hyperphosphorylation or C-55PP sequestering, but most likely to defective lipopolysaccharide transport. Indeed, lpxT overexpression in mutants lacking the L,D-transpeptidase LdtD and LdtE, which protect cells with outer membrane defects from osmotic lysis, caused cell envelope defects. Moreover, we found that pyrophosphorylated lipid A was also produced in a lpxT deletion mutant, indicating that LpxT is not the only protein able to perform such lipid A modification in E. coli.

scholarly journals Characterization of YmgF, a 72-Residue Inner Membrane Protein That Associates with the Escherichia coli Cell Division Machinery

2008 ◽  
Vol 191 (1) ◽  
pp. 333-346 ◽  
Author(s):  
Gouzel Karimova ◽  
Carine Robichon ◽  
Daniel Ladant
Keyword(s):  
Escherichia Coli ◽  
Cell Division ◽  
Membrane Protein ◽  
Integral Membrane Protein ◽  
Dependent Manner ◽  
E Coli ◽  
Division Site ◽  
Inner Membrane Protein ◽  
Two Hybrid

ABSTRACT Formation of the Escherichia coli division septum is catalyzed by a number of essential proteins (named Fts) that assemble into a ring-like structure at the future division site. Many of these Fts proteins are intrinsic transmembrane proteins whose functions are largely unknown. In the present study, we attempted to identify a novel putative component(s) of the E. coli cell division machinery by searching for proteins that could interact with known Fts proteins. To do that, we used a bacterial two-hybrid system based on interaction-mediated reconstitution of a cyclic AMP (cAMP) signaling cascade to perform a library screening in order to find putative partners of E. coli cell division protein FtsL. Here we report the characterization of YmgF, a 72-residue integral membrane protein of unknown function that was found to associate with many E. coli cell division proteins and to localize to the E. coli division septum in an FtsZ-, FtsA-, FtsQ-, and FtsN-dependent manner. Although YmgF was previously shown to be not essential for cell viability, we found that when overexpressed, YmgF was able to overcome the thermosensitive phenotype of the ftsQ1(Ts) mutation and restore its viability under low-osmolarity conditions. Our results suggest that YmgF might be a novel component of the E. coli cell division machinery.

scholarly journals Inner Membrane Protein YhcB Interacts with RodZ Involved in Cell Shape Maintenance in Escherichia coli

2012 ◽  
Vol 2012 ◽  
pp. 1-8 ◽  
Author(s):  
Gaochi Li ◽  
Kentaro Hamamoto ◽  
Madoka Kitakawa
Keyword(s):  
Escherichia Coli ◽  
Membrane Protein ◽  
Hybrid System ◽  
Cell Shape ◽  
Deletion Mutant ◽  
Cell Envelope ◽  
Inner Membrane ◽  
Inner Membrane Protein ◽  
Synthetically Lethal ◽  
Two Hybrid

Depletion of YhcB, an inner membrane protein of Escherichia coli, inhibited the growth of rodZ deletion mutant showing that the loss of both YhcB and RodZ is synthetically lethal. Furthermore, YhcB was demonstrated to interact with RodZ as well as several other proteins involved in cell shape maintenance and an inner membrane protein YciS of unknown function, using bacterial two-hybrid system. These observations seem to indicate that YhcB is involved in the biogenesis of cell envelope and the maintenance of cell shape together with RodZ.

Recruitment of the TolA protein to cell constriction sites in Escherichia coli via three separate mechanisms, and a critical role for FtsWI activity in recruitment of both TolA and TolQ.

2021 ◽  
Author(s):  
Cynthia A. Hale ◽  
Logan Persons ◽  
Piet A. J. de Boer
Keyword(s):  
Escherichia Coli ◽  
Cell Division ◽  
Cell Envelope ◽  
Critical Role ◽  
Gram Negative Bacteria ◽  
Bacterial Cells ◽  
Future Studies ◽  
E Coli ◽  
Middle Domain ◽  
Fundamental Biological Process

The Tol-Pal system of Gram-negative bacteria helps maintain integrity of the cell envelope and ensures that invagination of the envelope layers during cell fission occurs in a well-coordinated manner. In E. coli , the five Tol-Pal proteins (TolQ, R, A, B and Pal) accumulate at cell constriction sites in a manner that normally requires the activity of the cell constriction initiation protein FtsN. While septal recruitment of TolR, TolB and Pal also requires the presence of TolQ and/or TolA, each of the the latter two can recognize constriction sites independently of the other system proteins. What attracts TolQ or TolA to these sites is unclear. We show that FtsN attracts both proteins in an indirect fashion, and that PBP1A, PBP1B and CpoB are dispensable for their septal recruitment. However, the β-lactam aztreonam readily interferes with septal accumulation of both TolQ and TolA, indicating that FtsN-stimulated production of septal peptidoglycan by the FtsWI synthase is critical to their recruitment. We also discovered that each of TolA's three domains can recognize division sites in a separate fashion. Notably, the middle domain (TolAII) is responsible for directing TolA to constriction sites in the absence of other Tol-Pal proteins and CpoB, while recruitment of TolAI and TolAIII requires TolQ and a combination of TolB, Pal, and CpoB, respectively. Additionally, we describe the construction and use of functional fluorescent sandwich fusions of the ZipA division protein, which should be more broadly valuable in future studies of the E. coli cell division machinery. IMPORTANCE Cell division (cytokinesis) is a fundamental biological process that is incompletely understood for any organism. Division of bacterial cells relies on a ring-like machinery called the septal ring or divisome that assembles along the circumference of the mother cell at the site where constriction will eventually occur. In the well-studied bacterium Escherichia coli , this machinery contains over thirty distinct proteins. We studied how two such proteins, TolA and TolQ, which also play a role in maintaining integrity of the outer-membrane, are recruited to the machinery. We find that TolA can be recruited by three separate mechanisms, and that both proteins rely on the activity of a well-studied cell division enzyme for their recruitment.

Localization of the dicarboxylate binding protein in the cell envelope of Escherichia coli K12

1980 ◽  
Vol 58 (10) ◽  
pp. 885-897 ◽  
Author(s):  
Mary A. Bewick ◽  
Theodore C. Y. Lo
Keyword(s):  
Escherichia Coli ◽  
Cell Surface ◽  
Cell Envelope ◽  
Binding Protein ◽  
Osmotic Shock ◽  
Outer Region ◽  
Periplasmic Space ◽  
Whole Cells ◽  
Escherichia Coli K12 ◽  
The Moment

Examination of the localization of the dicarboxylate binding protein (DBP) in the cell envelope of Escherichia coli K12 reveals that this protein is present on the cell surface, and also in the inner and outer regions of the periplasmic space. The cell surface DBP is released by treating the cells with EDTA. This protein can be surface labeled by lactoperoxidase radio-iodination, and by diazo[125I]iodosulfanilic acid in whole cells. It also binds tightly, but not covalently, to lipopolysaccharide. The DBP located in the outer region of the periplasmic space is released when the outer membrane is dissociated by EDTA – osmotic shock treatment. The DBP located in the inner region of the periplasmic space is released only when the EDTA – osmotic shocked cells are subjected to lysozyme treatment. At the moment, it is not certain whether this protein is bound to or trapped by the peptidoglycan network. This protein cannot be surface labeled in whole cells or in EDTA – osmotic shock treated cells; and it is not associated with lipopolysaccharide. Analysis of transport mutants indicates that these DBP are coded by the same gene.

scholarly journals Bacterial Metal Resistance: Coping with Copper without Cooperativity?

mBio ◽  
2021 ◽  
Author(s):  
Nicholas P. Greene ◽  
Vassilis Koronakis
Keyword(s):  
Escherichia Coli ◽  
Cell Division ◽  
Cell Envelope ◽  
Metal Resistance ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Content Type ◽  
E Coli ◽  
Rnd Transporter ◽  
Entire Cell

In Escherichia coli and other Gram-negative bacteria, tripartite efflux pumps (TEPs) span the entire cell envelope and serve to remove noxious molecules from the cell. CusBCA is a TEP responsible for copper and silver detoxification in E. coli powered by the resistance-nodulation-cell division (RND) transporter, CusA.

scholarly journals SPOR Proteins Are Required for Functionality of Class A Penicillin-Binding Proteins in Escherichia coli

mBio ◽  
2020 ◽  
Vol 11 (6) ◽  
Author(s):  
Manuel Pazos ◽  
Katharina Peters ◽  
Adrien Boes ◽  
Yalda Safaei ◽  
Calem Kenward ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Cell Division ◽  
Binding Proteins ◽  
Cell Envelope ◽  
Synthetic Activity ◽  
Penicillin Binding Protein ◽  
Direct Role ◽  
Content Type ◽  
Class A

ABSTRACT Sporulation-related repeat (SPOR) domains are present in many bacterial cell envelope proteins and are known to bind peptidoglycan. Escherichia coli contains four SPOR proteins, DamX, DedD, FtsN, and RlpA, of which FtsN is essential for septal peptidoglycan synthesis. DamX and DedD may also play a role in cell division, based on mild cell division defects observed in strains lacking these SPOR domain proteins. Here, we show by nuclear magnetic resonance (NMR) spectroscopy that the periplasmic part of DedD consists of a disordered region followed by a canonical SPOR domain with a structure similar to that of the SPOR domains of FtsN, DamX, and RlpA. The absence of DamX or DedD decreases the functionality of the bifunctional transglycosylase-transpeptidase penicillin-binding protein 1B (PBP1B). DamX and DedD interact with PBP1B and stimulate its glycosyltransferase activity, and DamX also stimulates the transpeptidase activity. DedD also binds to PBP1A and stimulates its glycosyltransferase activity. Our data support a direct role of DamX and DedD in enhancing the activity of PBP1B and PBP1A, presumably during the synthesis of the cell division septum. IMPORTANCE Escherichia coli has four SPOR proteins that bind peptidoglycan, of which FtsN is essential for cell division. DamX and DedD are suggested to have semiredundant functions in cell division based on genetic evidence. Here, we solved the structure of the SPOR domain of DedD, and we show that both DamX and DedD interact with and stimulate the synthetic activity of the peptidoglycan synthases PBP1A and PBP1B, suggesting that these class A PBP enzymes act in concert with peptidoglycan-binding proteins during cell division.

scholarly journals YtfB, an OapA Domain-Containing Protein, Is a New Cell Division Protein in Escherichia coli

2018 ◽  
Vol 200 (13) ◽  
Author(s):  
Matthew A. Jorgenson ◽  
Kevin D. Young
Keyword(s):  
Escherichia Coli ◽  
Cell Wall ◽  
Cell Division ◽  
Wall Structure ◽  
Bacterial Cell Division ◽  
Negative Bacterium ◽  
Gram Negative ◽  
Content Type ◽  
Cell Division Protein ◽  
Inner Membrane Protein

ABSTRACT While screening the Pfam database for novel peptidoglycan (PG) binding modules, we identified the OapA domain, which is annotated as a LysM-like domain. LysM domains bind PG and mediate localization to the septal ring. In the Gram-negative bacterium Escherichia coli , an OapA domain is present in YtfB, an inner membrane protein of unknown function but whose overproduction causes cells to filament. Together, these observations suggested that YtfB directly affects cell division, most likely through its OapA domain. Here, we show that YtfB accumulates at the septal ring and that its action requires the division-initiating protein FtsZ and, to a lesser extent, ZipA, an early recruit to the septalsome. While the loss of YtfB had no discernible impact, a mutant lacking both YtfB and DedD (a known cell division protein) grew as filamentous cells. The YtfB OapA domain by itself also localized to sites of division, and this localization was enhanced by the presence of denuded PGs. Finally, the OapA domain bound PG, though binding did not depend on the formation of denuded glycans. Collectively, our findings demonstrate that YtfB is a cell division protein whose function is related to cell wall hydrolases. IMPORTANCE All living cells must divide in order to thrive. In bacteria, this involves the coordinated activities of a large number of proteins that work in concert to constrict the cell. Knowing which proteins contribute to this process and how they function is fundamental. Here, we identify a new member of the cell division apparatus in the Gram-negative bacterium Escherichia coli whose function is related to the generation of a transient cell wall structure. These findings deepen our understanding of bacterial cell division.

scholarly journals Role of Peptidoglycan Amidases in the Development and Morphology of the Division Septum in Escherichia coli

2007 ◽  
Vol 189 (14) ◽  
pp. 5334-5347 ◽  
Author(s):  
Richa Priyadarshini ◽  
Miguel A. de Pedro ◽  
Kevin D. Young
Keyword(s):  
Escherichia Coli ◽  
Cell Division ◽  
Cell Envelope ◽  
Penicillin Binding Protein ◽  
Mature Cell ◽  
Daughter Cells ◽  
Peptidoglycan Synthesis ◽  
A Cell ◽  
Entire Cell

ABSTRACT Escherichia coli contains multiple peptidoglycan-specific hydrolases, but their physiological purposes are poorly understood. Several mutants lacking combinations of hydrolases grow as chains of unseparated cells, indicating that these enzymes help cleave the septum to separate daughter cells after cell division. Here, we confirm previous observations that in the absence of two or more amidases, thickened and dark bands, which we term septal peptidoglycan (SP) rings, appear at division sites in isolated sacculi. The formation of SP rings depends on active cell division, and they apparently represent a cell division structure that accumulates because septal synthesis and hydrolysis are uncoupled. Even though septal constriction was incomplete, SP rings exhibited two properties of mature cell poles: they behaved as though composed of inert peptidoglycan, and they attracted the IcsA protein. Despite not being separated by a completed peptidoglycan wall, adjacent cells in these chains were often compartmentalized by the inner membrane, indicating that cytokinesis could occur in the absence of invagination of the entire cell envelope. Finally, deletion of penicillin-binding protein 5 from amidase mutants exacerbated the formation of twisted chains, producing numerous cells having septa with abnormal placements and geometries. The results suggest that the amidases are necessary for continued peptidoglycan synthesis during cell division, that their activities help create a septum having the appropriate geometry, and that they may contribute to the development of inert peptidoglycan.

Sign in / Sign up

Export Citation Format

Share Document