scholarly journals Mutant deletions in Escherichia coli affect the cellular levels of undecaprenyl phosphate and undecaprenyl diphosphate

2013 ◽  
Vol 13 (3) ◽  
pp. 86-91 ◽  
Author(s):  
Yuichiro Saito ◽  
Tatsuya Ishikawa ◽  
Marie Murakami ◽  
Keisuke Suzuki ◽  
Shingo Fujisaki
Keyword(s):  
Escherichia Coli ◽  
Undecaprenyl Phosphate

scholarly journals Interrupting Biosynthesis of O Antigen or the Lipopolysaccharide Core Produces Morphological Defects in Escherichia coli by Sequestering Undecaprenyl Phosphate

2016 ◽  
Vol 198 (22) ◽  
pp. 3070-3079 ◽  
Author(s):  
Matthew A. Jorgenson ◽  
Kevin D. Young
Keyword(s):  
Escherichia Coli ◽  
Cell Envelope ◽  
Negative Bacterium ◽  
Core Oligosaccharide ◽  
Gram Negative ◽  
Content Type ◽  
O Antigen ◽  
Dead End ◽  
Morphological Defects ◽  
Undecaprenyl Phosphate

ABSTRACTUndecaprenyl phosphate (Und-P) is a member of the family of essential polyprenyl phosphate lipid carriers and in the Gram-negative bacteriumEscherichia coliis required for synthesizing the peptidoglycan (PG) cell wall, enterobacterial common antigen (ECA), O antigen, and colanic acid. Previously, we found that interruption of ECA biosynthesis indirectly alters PG synthesis by sequestering Und-P via dead-end intermediates, causing morphological defects. To determine if competition for Und-P was a more general phenomenon, we determined if O-antigen intermediates caused similar effects. Indeed, disrupting the synthesis of O antigen or the lipopolysaccharide core oligosaccharide induced cell shape deformities, which were suppressed by preventing the initiation of O-antigen biosynthesis or by manipulating Und-P metabolism. We conclude that accumulation of O-antigen intermediates alters PG synthesis by sequestering Und-P. Importantly, many previous experiments addressed the physiological functions of various oligosaccharides and glycoconjugates, but these studies employed mutants that accumulate deleterious intermediates. Thus, conclusions based on these experiments must be reevaluated to account for possible indirect effects of Und-P sequestration.IMPORTANCEBacteria use long-chain isoprenoids like undecaprenyl phosphate (Und-P) as lipid carriers to assemble numerous glycan polymers that comprise the cell envelope. In any one bacterium, multiple oligosaccharide biosynthetic pathways compete for a common pool of Und-P, which means that disruptions in one pathway may produce secondary consequences that affect the others. Using the Gram-negative bacteriumEscherichia colias a model, we demonstrate that interruption of the biogenesis of O antigen, a major outer membrane component, indirectly impairs peptidoglycan synthesis by sequestering Und-P into dead-end intermediates. These results strongly argue that the functions of many Und-P-utilizing pathways must be reevaluated, because much of our current understanding is based on experiments that did not control for these unintended secondary effects.

scholarly journals A genetic screen to identify factors affected by undecaprenyl phosphate recycling uncovers novel connections to morphogenesis in Escherichia coli

2020 ◽  
Author(s):  
Matthew A. Jorgenson ◽  
Joseph C. Bryant
Keyword(s):  
Escherichia Coli ◽  
Cell Wall ◽  
Cytoplasmic Membrane ◽  
Response Regulator ◽  
Genetic Screen ◽  
Glutathione Metabolism ◽  
Cell Integrity ◽  
Two Component System ◽  
Dna Replication And Repair ◽  
Undecaprenyl Phosphate

AbstractUndecaprenyl phosphate (Und-P) is an essential lipid carrier that ferries cell wall intermediates across the cytoplasmic membrane in bacteria. Und-P is generated by dephosphorylating undecaprenyl diphosphate (Und-PP). In Escherichia coli, BacA, PgpB, YbjG, and LpxT dephosphorylate Und-PP and are conditionally essential. To identify vulnerabilities that arise when Und-P metabolism is defective, we developed a genetic screen for synthetic mutations in combination with ΔybjG ΔlpxT ΔbacA. The screen uncovered system-wide connections, including novel connections to cell division, DNA replication and repair, signal transduction, and glutathione metabolism. Further analysis revealed several new morphogenes; loss of one of these, qseC, caused cells to enlarge and lyse. QseC is the sensor kinase component of the QseBC two-component system. In the absence of QseC, the QseB response regulator is overactivated by PmrB cross-phosphorylation. Here, we show that deleting qseB completely reverses the shape defect of ΔqseC cells, as does overexpressing rprA (a small RNA). Surprisingly, deleting pmrB only partially suppressed qseC-related shape defects. Thus, QseB is activated by multiple factors in the absence of QseC and functions ascribed to QseBC may be related to cell wall defects. Altogether, our findings provide a framework for identifying new determinants of cell integrity that could be targeted in future therapies.

scholarly journals An Undecaprenyl Phosphate-Aminoarabinose Flippase Required for Polymyxin Resistance in Escherichia coli

2007 ◽  
Vol 282 (49) ◽  
pp. 36077-36089 ◽  
Author(s):  
Aixin Yan ◽  
Ziqiang Guan ◽  
Christian R. H. Raetz
Keyword(s):  
Escherichia Coli ◽  
Transcription Factor ◽  
Antimicrobial Peptides ◽  
Salmonella Typhimurium ◽  
Lipid A ◽  
Inner Membrane ◽  
Cationic Antimicrobial Peptides ◽  
E Coli ◽  
Undecaprenyl Phosphate

Modification of lipid A with the 4-amino-4-deoxy-l-arabinose (l-Ara4N) moiety is required for resistance to polymyxin and cationic antimicrobial peptides in Escherichia coli and Salmonella typhimurium. An operon of seven genes (designated pmrHFIJKLM in S. typhimurium), which is regulated by the PmrA transcription factor and is also present in E. coli, is necessary for the maintenance of polymyxin resistance. We previously elucidated the roles of pmrHFIJK in the biosynthesis and attachment of l-Ara4N to lipid A and renamed these genes arn-BCADT, respectively. We now propose functions for the last two genes of the operon, pmrL and pmrM. Chromosomal inactivation of each of these genes in an E. coli pmrAc parent switched its phenotype from polymyxin-resistant to polymyxin-sensitive. Lipid A was no longer modified with l-Ara4N, even though the levels of the lipid-linked donor of the l-Ara4N moiety, undecaprenyl phosphate-α-l-Ara4N, were not reduced in the mutants. However, the undecaprenyl phosphate-α-l-Ara4N present in the mutants was less concentrated on the periplasmic surface of the inner membrane, as judged by 4-5-fold reduced labeling with the inner membrane-impermeable amine reagent N-hydroxysulfosuccin-imidobiotin. In an arnT mutant of the same pmrAc parent, which lacks the enzyme that transfers the l-Ara4N unit to lipid A but retains the same high levels of undecaprenyl phosphate-α-l-Ara4N as the parent, N-hydroxysulfosuccinimidobiotin labeling was not reduced. These results implicate pmrL and pmrM, but not arnT, in transporting undecaprenyl phosphate-α-l-Ara4N across the inner membrane. PmrM and PmrL, now renamed ArnE and ArnF because of their involvement in l-Ara4N modification of lipid A, may be subunits of an undecaprenyl phosphate-α-l-Ara4N flippase.

scholarly journals An Escherichia coli undecaprenyl-pyrophosphate phosphatase implicated in undecaprenyl phosphate recycling

Microbiology ◽  
2007 ◽  
Vol 153 (8) ◽  
pp. 2518-2529 ◽  
Author(s):  
Laura D. Tatar ◽  
Cristina L. Marolda ◽  
Andrew N. Polischuk ◽  
Deborah van Leeuwen ◽  
Miguel A. Valvano
Keyword(s):  
Escherichia Coli ◽  
Undecaprenyl Phosphate

scholarly journals CbrA Mediates Colicin M Resistance in Escherichia coli through Modification of Undecaprenyl-Phosphate-Linked Peptidoglycan Precursors

2020 ◽  
Vol 202 (23) ◽  
Author(s):  
Hélène Barreteau ◽  
Delphine Patin ◽  
Ahmed Bouhss ◽  
Didier Blanot ◽  
Dominique Mengin-Lecreulx ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Cleavage Site ◽  
Resistance Mechanism ◽  
Content Type ◽  
E Coli ◽  
Lipid Ii ◽  
Colicin M ◽  
Undecaprenyl Phosphate

ABSTRACT Colicin M is an enzymatic bacteriocin produced by some Escherichia coli strains which provokes cell lysis of competitor strains by hydrolysis of the cell wall peptidoglycan undecaprenyl-PP-MurNAc(-pentapeptide)-GlcNAc (lipid II) precursor. The overexpression of a gene, cbrA (formerly yidS), was shown to protect E. coli cells from the deleterious effects of this colicin, but the underlying resistance mechanism was not established. We report here that a major structural modification of the undecaprenyl-phosphate carrier lipid and of its derivatives occurred in membranes of CbrA-overexpressing cells, which explains the acquisition of resistance toward this bacteriocin. Indeed, a main fraction of these lipids, including the lipid II peptidoglycan precursor, now displayed a saturated isoprene unit at the α-position, i.e., the unit closest to the colicin M cleavage site. Only unsaturated forms of these lipids were normally detectable in wild-type cells. In vitro and in vivo assays showed that colicin M did not hydrolyze α-saturated lipid II, clearly identifying this substrate modification as the resistance mechanism. These saturated forms of undecaprenyl-phosphate and lipid II remained substrates of the different enzymes participating in peptidoglycan biosynthesis and carrier lipid recycling, allowing this colicin M-resistance mechanism to occur without affecting this essential pathway. IMPORTANCE Overexpression of the chromosomal cbrA gene allows E. coli to resist colicin M (ColM), a bacteriocin specifically hydrolyzing the undecaprenyl-PP-MurNAc(-pentapeptide)-GlcNAc (lipid II) peptidoglycan precursor of targeted cells. This resistance results from a CbrA-dependent modification of the precursor structure, i.e., reduction of the α-isoprenyl bond of C55-carrier lipid moiety that is proximal to ColM cleavage site. This modification, observed here for the first time in eubacteria, annihilates the ColM activity without affecting peptidoglycan biogenesis. These data, which further increase our knowledge of the substrate specificity of this colicin, highlight the capability of E. coli to generate reduced forms of C55-carrier lipid and its derivatives. Whether the function of this modification is only relevant with respect to ColM resistance is now questioned.

Structural organization of escherichia coli ribosomes as determined by immuno electron microscopy

1978 ◽  
Vol 36 (2) ◽  
pp. 166-167 ◽  
Author(s):  
G. Stöffler ◽  
R.W. Bald ◽  
J. Dieckhoff ◽  
H. Eckhard ◽  
R. Lührmann ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Escherichia Coli ◽  
Ribosomal Proteins ◽  
Structural Organization ◽  
Three Dimensional ◽  
Ribosomal Subunit ◽  
Spatial Arrangement ◽  
Small Subunit ◽  
Antibody Binding ◽  
Immuno Electron Microscopy

A central step towards an understanding of the structure and function of the Escherichia coli ribosome, a large multicomponent assembly, is the elucidation of the spatial arrangement of its 54 proteins and its three rRNA molecules. The structural organization of ribosomal components has been investigated by a number of experimental approaches. Specific antibodies directed against each of the 54 ribosomal proteins of Escherichia coli have been performed to examine antibody-subunit complexes by electron microscopy. The position of the bound antibody, specific for a particular protein, can be determined; it indicates the location of the corresponding protein on the ribosomal surface.The three-dimensional distribution of each of the 21 small subunit proteins on the ribosomal surface has been determined by immuno electron microscopy: the 21 proteins have been found exposed with altogether 43 antibody binding sites. Each one of 12 proteins showed antibody binding at remote positions on the subunit surface, indicating highly extended conformations of the proteins concerned within the 30S ribosomal subunit; the remaining proteins are, however, not necessarily globular in shape (Fig. 1).

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