Structural characterization of the essential cell division protein FtsE and its interaction with FtsX in Streptococcus pneumoniae
TL;DR: Structural analysis reveals that FtsE contains all the conserved structural motifs associated with ATPase activity, and allowed interpretation of the in vivo dimeric arrangement in both ADP and ATP states, and provides evidences on how difference between ATP and ADP states in FTSE would dramatically alter FtsEX interaction with PG hydrolase PcsB in pneumococcal division.
Abstract: FtsEX is a membrane complex widely conserved across diverse bacterial genera and involved in critical processes such as recruitment of division proteins and in spatial and temporal regulation of muralytic activity during cell division or sporulation. FtsEX is a member of the ABC transporter superfamily, where FtsX is an integral membrane protein and FtsE is an ATPase, required for mechanotransmission of the signal from the cytosol through the membrane, to regulate the activity of cell-wall hydrolases in the periplasm. Both proteins are essential in the major human respiratory pathogenic bacterium, Streptococcus pneumoniae and interact with the modular peptidoglycan hydrolase PcsB at the septum. Here, we report the high-resolution structures of pneumococcal FtsE in complex with different nucleotides. Structural analysis reveals that FtsE contains all the conserved structural motifs associated with ATPase activity, and allowed interpretation of the in vivo dimeric arrangement in both ADP and ATP states. Interestingly, three specific FtsE regions were identified with high structural plasticity that shape the cavity in which the cytosolic region of FtsX would be inserted. The residues corresponding to the FtsX coupling helix, responsible for FtsE contact, were identified and validated by in vivo mutagenesis studies showing that this interaction is essential for cell growth and proper morphology. IMPORTANCE Bacterial cell division is a central process that requires exquisite orchestration of both the cell wall biosynthetic and lytic machineries. The essential membrane complex FtsEX, widely conserved across bacteria, play a central role by recruiting proteins to the divisome apparatus and by regulating periplasmic muralytic activity from the cytosol. FtsEX is a member of the Type VII family of the ABC-superfamily but instead transporter, couple ATP hydrolysis by FtsE to mechanically transduce a conformational signal to activate PG hydrolases. So far, no structural information is available for FtsE. Here we provide the structural characterization of FtsE confirming its ATPase nature and revealing regions with high structural plasticity key for FtsX binding. The complementary region in FtsX has been also identified and validated in vivo. Our results provide evidences on how difference between ATP and ADP states in FtsE would dramatically alter FtsEX interaction with PG hydrolase PcsB in pneumococcal division.
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01 Jan 2004
TL;DR: A large class of ATPases contain a RecA-like structural domain and uses the energy of nucleotide binding and hydrolysis to perform mechanical work to move polypeptides or nucleic acids as discussed by the authors.
Abstract: A large class of ATPases contains a RecA-like structural domain and uses the energy of nucleotide binding and hydrolysis to perform mechanical work, for example, to move polypeptides or nucleic acids. These ATPases include helicases, ABC transporters, clamp loaders, and proteases. The functional units of the ATPases contain different numbers of RecA-like domains, but the nucleotide is always bound at the interface between two adjacent RecA-like folds and the two domains move relative to one another during the ATPase cycle. The structures determined for different RecA-like motor ATPases begin to reveal how they move macromolecules.
140 citations
TL;DR: Together, these results reveal new aspects of spatially ordered PG synthesis in ovococcal bacteria during cell division.
Abstract: Bacterial peptidoglycan (PG) synthesis requires strict spatiotemporal organization to reproduce specific cell shapes. In ovoid-shaped Streptococcus pneumoniae (Spn), septal and peripheral (elongation) PG synthesis occur simultaneously at midcell. To uncover the organization of proteins and activities that carry out these two modes of PG synthesis, we examined Spn cells vertically oriented onto their poles to image the division plane at the high lateral resolution of 3D-SIM (structured-illumination microscopy). Labeling with fluorescent D-amino acids (FDAA) showed that areas of new transpeptidase (TP) activity catalyzed by penicillin-binding proteins (PBPs) separate into a pair of concentric rings early in division, representing peripheral PG (pPG) synthesis (outer ring) and the leading-edge (inner ring) of septal PG (sPG) synthesis. Fluorescently tagged PBP2x or FtsZ locate primarily to the inner FDAA-marked ring, whereas PBP2b and FtsX remain in the outer ring, suggesting roles in sPG or pPG synthesis, respectively. Pulses of FDAA labeling revealed an arrangement of separate regularly spaced "nodes" of TP activity around the division site of predivisional cells. Tagged PBP2x, PBP2b, and FtsX proteins also exhibited nodal patterns with spacing comparable to that of FDAA labeling. Together, these results reveal new aspects of spatially ordered PG synthesis in ovococcal bacteria during cell division.
20 citations
Cites background or result from "Structural characterization of the ..."
...FtsX is a polytopic membrane protein that forms a complex with the cytoplasmic FtsE ATPase and the extracellular PcsB PG hydrolase in Spn (Alcorlo et al., 2020; Rued et al., 2019; Sham et al., 2011, 2013)....
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...FtsX is an essential, polytopic membrane protein that forms a complex with the cytoplasmic FtsE ATPase and the extracellular PcsB PG hydrolase in Spn (Alcorlo et al., 2020, Bartual et al., 2014, Rued et al., 2019, Sham et al., 2011, 2013)....
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...The FtsEX:PcsB complex is essential for Spn growth due to its role in hydrolytic remodeling during PG synthesis (Alcorlo et al., 2020, Bartual et al., 2014, Ng et al., 2004)....
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...Depletion of essential FtsX, FtsE, or PcsB results in chains of spherical cells, similar to those caused by depletion of bPBP2b (Alcorlo et al., 2020, Berg et al., 2013, Ng et al., 2004, Sham et al., 2011, 2013, Tsui et al., 2014)....
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...Depletion of FtsEX:PcsB results in the formation of chains of spheroid cells (Ng et al., 2004, Sham et al., 2013), characteristic of defective pPG synthesis (Alcorlo et al., 2020, Tsui et al., 2014)....
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TL;DR: In this article , the structure of FtsEX in complex with an amidase and amidase activator is reported, revealing how ATP binding to the complex promotes amidase activation and providing structural information that may help target the activation mechanism for the development of cell lysis inducing antibiotics.
Abstract: Significance Bacterial division is an essential cellular process that involves the formation of a septum made of peptidoglycan. The septum is initially shared between daughters and must be processed to complete division. Septal splitting has long been known to be mediated by enzymes called amidases that are controlled by an activator protein and the ABC-transporter-like complex called FtsEX. However, the mechanism of amidase regulation by this system has remained unclear. Here, we report the structure of FtsEX in complex with an amidase and amidase activator, revealing how ATP binding to the complex promotes amidase activation and providing structural information that may help target the activation mechanism for the development of cell lysis inducing antibiotics.
1 citations
TL;DR: There is distinct spatial separation of the sPG machine, including FtsZ, from the pPG synthesis machine at the midcell of dividing Spn cells, and in predivisional cells, PBPs and TP activity are organized heterogeneously into regularly spaced nodes, whose number and dynamic distribution are likely driven by the PG synthesis of PBP:SEDS complexes.
Abstract: Bacterial peptidoglycan (PG) synthesis requires strict spatial and temporal organization to reproduce specific cell shapes. In the ovoid-shaped, pathogenic bacterium Streptococcus pneumoniae (Spn), septal and peripheral (sidewall-like) PG synthesis occur simultaneously at midcell. To uncover the organization of proteins and activities that carry out these two modes of PG synthesis, we examined Spn cells vertically oriented onto their poles to image the division plane at the high lateral resolution of 3D-SIM (structured-illumination microscopy). Using fluorescent D-amino acid (FDAA) probes, we show that areas of new transpeptidase (TP) activity catalyzed by penicillin-binding proteins (PBPs) separate into a pair of concentric rings early in division, representing peripheral PG (pPG) synthesis (outer ring) and the leading-edge (inner ring) of septal PG (sPG) synthesis. Fluorescently tagged PBP2x or FtsZ locate primarily to the inner FDAA-marked ring, whereas PBP2b and FtsX remain in the outer ring, suggesting roles in sPG or pPG synthesis, respectively. Short pulses of FDAA labeling revealed an arrangement of separate regularly spaced nodes of TP activity around the division site of predivisional cells. Control experiments in wild-type and mutant strains support the interpretation of nodal spacing of TP activity, and statistical analysis confirmed that the number of nodes correlates with different ring diameters. This nodal pattern of FDAA labeling is conserved in other ovoid-shaped species. Tagged PBP2x, PBP2b, and FtsX proteins also exhibited nodal patterns with spacing comparable to that of FDAA labeling. Together, these results reveal a highly ordered PG synthesis apparatus in ovococcal bacteria at different stages of division.
1 citations
Cites background from "Structural characterization of the ..."
...The FtsEX:PcsB complex is essential 230 for Spn growth due to its role in hydrolytic remodeling during PG synthesis (28, 32, 33)....
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...11 Depletion of FtsEX:PcsB results in the formation of chains of spheroid cells (31, 33), 232 characteristic of defective pPG synthesis (11, 28)....
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...the extracellular PcsB PG hydrolase in Spn (28-32)....
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...polytopic membrane protein that forms a complex with the cytoplasmic FtsE ATPase and 229 the extracellular PcsB PG hydrolase in Spn (28-31)....
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TL;DR: In this paper , two saline-alkali-tolerant bacterial strains, Priestia aryabhattai JL-5 and Staphylococcus pseudoxylosus XW-4, were isolated, with high capabilities of hydrolyzing phosphate and producing cellulase, respectively.
Abstract: Two saline-alkali-tolerant bacterial strains, Priestia aryabhattai JL-5 and Staphylococcus pseudoxylosus XW-4, were isolated, with high capabilities of hydrolyzing phosphate and producing cellulase, respectively. The molecular mechanisms regulating the saline-alkali tolerance in the strain JL-5 were further investigated using transcriptome analysis. The contents of lactic acid and proline and the enzymatic activity of glutamine synthetase in the strain JL-5 were significantly increased. The properties of saline-alkali soils were significantly improved by the enhanced growth of the indicator plant Leymus chinensis under the combined applications of the strains JL-5 and XW-4 mixed with corn straw. The contents of catalase, peroxidase, superoxide dismutase and proline of L. chinensis were significantly increased, and the content of malondialdehyde was significantly decreased in the combined treatment of both bacterial strains. The contents of available nitrogen, phosphorus and potassium and organic matters in the soil treated with both strains were significantly increased, as well as the diversity and abundance of the soil microbiota. Our study evidently demonstrated the synergistic effects of the strains JL-5 and XW-4, indicator plants and the local microbiota in terms of improving the saline-alkali soil properties, providing strong experimental evidence to support the commercial development of the combined application of both strains to improve the properties of saline-alkali soils.
References
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TL;DR: Coot is a molecular-graphics program designed to assist in the building of protein and other macromolecular models and the current state of development and available features are presented.
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TL;DR: An analysis of the solvent content of 116 different crystal forms of globular proteins found that in many cases this range will be sufficiently restrictive to enable the probable number of molecules in the crystallographic asymmetric unit to be determined directly from the molecular weight of the protein and the space group and unit cell dimensions of the crystal.
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TL;DR: An update to the SWISS-MODEL server is presented, which includes the implementation of a new modelling engine, ProMod3, and the introduction a new local model quality estimation method, QMEANDisCo.
Abstract: Homology modelling has matured into an important technique in structural biology, significantly contributing to narrowing the gap between known protein sequences and experimentally determined structures. Fully automated workflows and servers simplify and streamline the homology modelling process, also allowing users without a specific computational expertise to generate reliable protein models and have easy access to modelling results, their visualization and interpretation. Here, we present an update to the SWISS-MODEL server, which pioneered the field of automated modelling 25 years ago and been continuously further developed. Recently, its functionality has been extended to the modelling of homo- and heteromeric complexes. Starting from the amino acid sequences of the interacting proteins, both the stoichiometry and the overall structure of the complex are inferred by homology modelling. Other major improvements include the implementation of a new modelling engine, ProMod3 and the introduction a new local model quality estimation method, QMEANDisCo. SWISS-MODEL is freely available at https://swissmodel.expasy.org.
7,022 citations
"Structural characterization of the ..." refers methods in this paper
...All our attempts to co-crystallize FtsE with this FtsX fragment failed, but a three-dimensional model for this sequence was generated with the Swiss-Model server (47) that identified PDB 5NIK as the best template sharing 32....
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