Bacterial outer membrane constriction

Egan, Alexander J F

doi:10.1111/mmi.13908

Cited by 92 publications

(81 citation statements)

References 108 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The integrity of these membranes is mainly governed by lipid–lipid, protein–lipid, and protein–protein interactions . These membranes have diverse lipid compositions and are active participants in essential biological processes . The outer membrane of Gram‐negative bacteria is the first of the three layers that surround the bacterial cytoplasm .…”

Section: Introductionmentioning

confidence: 99%

“…10 These membranes have diverse lipid compositions and are active participants in essential biological processes. 11 The outer membrane of Gram-negative bacteria is the first of the three layers that surround the bacterial cytoplasm. 12 This bilayer is highly asymmetric: the inner leaflet is mainly built by various species of phospholipids, while the outer leaflet is largely composed of lipopolysaccharides (LPS).…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

A simple protocol to characterize bacterial cell‐envelope lipoproteins in a native‐like environment

2019

View full text Add to dashboard Cite

Physiological conditions in living cells are strictly regulated to allow, optimize, and coordinate biological processes. The bacterial cell envelope is the compartment where the communication with the external environment takes place. This involves membrane proteins, key players in many biological processes that ensure bacterial survival. The biochemical characterization of membrane proteins, either integral, lipidated or peripheral is challenging due to their mixed protein‐lipid nature, making it difficult to purify and obtain considerable amounts of samples. In contrast to integral membrane proteins, lipidated proteins are usually purified as truncated soluble versions, neglecting the impact of the membrane environment. Here we report a simple and robust protocol to characterize bacterial lipidated proteins in spheroplasts from Escherichia coli using a β‐lactamase as a model. The Metallo‐β‐lactamase NDM‐1 is an enzyme anchored to the inner leaflet of the outer membrane of Gram‐negative bacteria. Kinetic parameters and stability of the lipidated NDM‐1 and the soluble unbound version (NDM‐1 C26A) were measured in spheroplasts and periplasm, respectively. These studies revealed that membrane anchoring increases the KM of the enzyme, consequently decreasing the catalytic efficiency, while not affecting its kinetic stability. This approach can be used to characterize lipidated proteins avoiding the purification step while mimicking its native environment. This approach also helps in filling the gap between in vitro and in vivo studies.

show abstract

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

A simple protocol to characterize bacterial cell‐envelope lipoproteins in a native‐like environment

2019

View full text Add to dashboard Cite

show abstract

“…Cycles of septal peptidoglycan synthesis/hydrolysis by the divisome and FtsZ treadmilling drive constriction of the entire cell envelope 2 . While much is known about the components that constrict the inner membrane and remodel the cell wall during cell division relatively little is known about how the outer membrane (OM) is invaginated 3 . The OM is essential in most Gram-negative bacteria 4,5 ; where it serves as both a permeability barrier to exclude hydrophobic and hydrophilic compounds, including antibiotics such as vancomycin 6 , and contributes to the mechanical stiffness of the cell 4 .…”

Section: Introductionmentioning

confidence: 99%

“…The Tol assembly is essential in bacteria expressing O-antigens, is a virulence factor in host-pathogen interactions 12-14 and is implicated in biofilm formation 15 . The core components of Tol are three IM proteins, TolQ, TolR and TolA, periplasmic TolB and peptidoglycan associated lipoprotein Pal in the inner leaflet of the OM ( Figure 1a ) 3 . TolA in the inner membrane spans the periplasm and undergoes PMF-driven conformational changes by virtue of its interaction partners TolQ and TolR, which are homologues of the MotA and MotB stator proteins that drive rotation of the bacterial flagellum ( Supplementary Figure 1 ) 16 .…”

Section: Introductionmentioning

confidence: 99%

Pal stabilises the bacterial outer membrane during constriction by a mobilisation-and-capture mechanism

Szczepaniak

Holmes

Rajasekar

et al. 2019

Preprint

View full text Add to dashboard Cite

25Coordination of outer membrane constriction with septation is critical to faithful division 26 in Gram-negative bacteria and vital to the barrier function of the membrane. Recent 27 studies suggest this coordination is through the active accumulation of the 28 peptidoglycan-binding outer membrane lipoprotein Pal at division sites by the Tol 29 system, but the mechanism is unknown. Here, we show that Pal accumulation at 30Escherichia coli division sites is a consequence of three key functions of the Tol system. 31First, Tol mobilises Pal molecules in dividing cells, which otherwise diffuse very slowly 32 due to their binding of the cell wall. Second, Tol actively captures mobilised Pal 33 molecules and deposits them at the division septum. Third, the active capture 34 mechanism is analogous to that used by the inner membrane protein TonB to dislodge 35 the plug domains of outer membrane TonB-dependent nutrient transporters. We 36 conclude that outer membrane constriction is coordinated with cell division by active 37 mobilisation-and-capture of Pal at division septa by the Tol system. 38 39 Word count, 160 40 41 42 43 44 45 46 47 across the inner membrane 7-9 . Recently, Petiti et al have suggested that the 60multiprotein Tol system (also known as Tol-Pal) constricts the OM by populating the 61 division septum with Pal 10 . However, no mechanism has been proposed. In the present 62 work, through a combination of in vivo imaging, deletion analysis and mutagenesis, 63 structure determination, biophysical measurements, mathematical modelling and 64 molecular dynamics simulations, we demonstrate that the PMF is exploited by the Tol 65 system to both mobilise Pal in the OM of dividing cells and to then capture these 66 mobilised molecules at division sites. Mobilisation-and-capture circumvents Pal's 67 intrinsically low mobility in the OM and results in its accumulation at division sites, where 68 it invaginates the OM through non-covalent interactions with newly-formed septal 69 peptidoglycan. 70 4 tol genes, which are found in most Gram-negative bacteria, were originally 71 identified by Luria and co-workers in the 1960s through mutations that engendered 72Escherichia coli tolerance towards colicins and filamentous bacteriophages 11 . 73Concomittant with this tolerance is a pleiotropic OM instability phenotype that is manifest 74 through cell filamentation and division defects, hypersensitivity towards detergents and 75 bile salts and leakage of periplasmic contents. The Tol assembly is essential in bacteria 76expressing O-antigens, is a virulence factor in host-pathogen interactions 12-14 and is 77 implicated in biofilm formation 15 . The core components of Tol are three IM proteins, 78TolQ, TolR and TolA, periplasmic TolB and peptidoglycan associated lipoprotein Pal in 79 the inner leaflet of the OM (Figure 1a) 3 . TolA in the inner membrane spans the 80 periplasm and undergoes PMF-driven conformational changes by virtue of its interaction 81 partners TolQ and TolR, which are homologues of the MotA and MotB stato...

show abstract

“…Functioning as a recruitment base, the protoring components then enlist the remaining essential division proteins to form a mature ‘divisome’ 5 . As soon as it is fully assembled, the divisome starts to constrict the cell envelope by reshaping the septal geometry, ultimately leading to sequential closure of the inner and outer membranes 8-10 .…”

Section: Introductionmentioning

confidence: 99%

Cell shape independent FtsZ dynamics in synthetically remodeled cells

Söderström¹,

Badrutdinov

Chan³

et al. 2018

Preprint

View full text Add to dashboard Cite

46The FtsZ protein is a key regulator of bacterial cell division. It has been implicated in 47 acting as a scaffolding protein for other division proteins, being a force generator 48 during constriction, and more recently, as an active regulator of septal cell wall 49 production. During an early stage of the division cycle, FtsZ assembles into a 50 heterogeneous structure coined the "Z-ring" due to its resemblance to a ring confined 51 by the midcell geometry. While in vitro experiments on supported lipid bilayers have 52 shown that purified FtsZ can self-organize into a swirling ring roughly the diameter of 53 a bacterial cell, it is not known how, and if, membrane curvature affects FtsZ assembly 54 and dynamics in vivo. 55To establish a framework for examining geometrical influences on proper Z-ring 56 assembly and dynamics, we sculptured Escherichia coli cells into unnatural shapes, 57 such as squares and hearts, using division-and cell wall-specific inhibitors in a micro 58 fabrication scheme. This approach allowed us to examine FtsZ behavior in engineered 59 "Z-squares" and "Z-hearts", and in giant cells up to 50 times their normal volume. 60 Quantification of super-resolution STimulated Emission Depletion (STED) nanoscopy 61data showed that FtsZ densities in sculptured cells maintained the same dimensions 62 as their wild-type counterparts. Additionally, time-resolved fluorescence 63 measurements revealed that FtsZ dynamics were generally conserved in a wide range 64 of cell shapes. Based on our results, we conclude that the underlying membrane 65 environment is not a deciding factor for FtsZ filament maintenance and treadmilling in 66 vivo. 67 68 Introduction 69Most bacterial cells divide by binary fission, whereby one mother cell splits into two 70 identical daughters 1-3 . Decades of study have led to a detailed understanding of how 71 the cell division machinery, the divisome, carries out this task during the later stages 72 of the cell cycle 4,5 . At the heart of this process is the eukaryotic tubulin homologue, 73FtsZ 6 that, together with its membrane anchors ZipA and FtsA, forms an intermediate 74 structure called the proto-ring ( Fig. 1a) 7 . Functioning as a recruitment base, the proto-75 ring components then enlist the remaining essential division proteins to form a mature 76 'divisome' 5 . As soon as it is fully assembled, the divisome starts to constrict the cell 77 envelope by reshaping the septal geometry, ultimately leading to sequential closure of 78 the inner and outer membranes [8][9][10] . 79In rod-shaped model bacteria such as Escherichia coli and Bacillus subtilis, FtsZ is 80 believed to organize into short bundles of filaments, roughly 100 nm in length 11,12 , that 81 treadmill at the septum with a circumferential velocity in the order of 20-30 nm/s [13][14][15] . 82The treadmilling filaments guide and regulate septal peptidoglycan (PG-) production 83 and ingrowth, leading up to septation 16 . This mode of action may be limited to rod-84 shaped bacteria that have two separate PG-mach...

show abstract

Bacterial outer membrane constriction

Cited by 92 publications

References 108 publications

A simple protocol to characterize bacterial cell‐envelope lipoproteins in a native‐like environment

A simple protocol to characterize bacterial cell‐envelope lipoproteins in a native‐like environment

Pal stabilises the bacterial outer membrane during constriction by a mobilisation-and-capture mechanism

Cell shape independent FtsZ dynamics in synthetically remodeled cells

Contact Info

Product

Resources

About