Characterization of CrgA, a New Partner of the Mycobacterium tuberculosis Peptidoglycan Polymerization Complexes

Płociński, Przemysław; Ziolkiewicz, M.; Kiran, Manjot; Vadrevu, Suryakiran; Nguyen, Hau B.; Hugonnet, Jean-Emmanuel; Veckerlé, Carole; Arthur, Michel; Dziadek, Jarosław; Cross, Timothy A.; Madiraju, Murty V. V. S.; Rajagopalan, Malini

doi:10.1128/jb.00188-11

Cited by 61 publications

(82 citation statements)

References 54 publications

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“…S3) including resonances from the disordered regions of the protein and the structured interfacial regions. Although a bacterial two-hybrid assay suggested that CrgA forms a dimer (1), the single resonances observed here for all of the residues indicate a monomer (or symmetric dimer) in these preparations. Fig.…”

Section: Resultsmentioning

confidence: 96%

“…CrgA has the responsibility of coordinating the assembly of multiple membrane proteins for cell division (1). CrgA binds to FtsZ, the protein responsible for initiating the formation of the Z-ring when cells divide.…”

Section: Discussionmentioning

confidence: 99%

“…In particular, FtsI, with a single TM helix, is a transpeptidase responsible for synthesis of the septal PG (1). A crgA-deletion mutant results in the loss of septal and polar localization of FtsI, suggesting the importance of CrgA for PG synthesis through its recruitment of FtsI.…”

Section: Significancementioning

confidence: 99%

“…CrgA, a transmembrane (TM) protein, is a central component of the Mtb divisome (1). CrgA has homologs in other actinomycetes (2,3), but not in the two bacteria, Escherichia coli and Bacillus subtilis, with better characterized cell division mechanisms.…”

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confidence: 99%

“…CrgA is localized at the poles and septum, and interacts with multiple cell division proteins, including FtsZ, FtsQ, FtsI (PBPB), PBPA, and CwsA. One function of these interactions is to stabilize the divisome (1,4). The interaction with CwsA, a protein that is unique to mycobacteria (5), might coordinate elongation at the poles and division at midcell (4).…”

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confidence: 99%

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Structure of CrgA, a cell division structural and regulatory protein fromMycobacterium tuberculosis, in lipid bilayers

Das

Dai

Hung

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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The 93-residue transmembrane protein CrgA in Mycobacterium tuberculosis is a central component of the divisome, a large macromolecular machine responsible for cell division. Through interactions with multiple other components including FtsZ, FtsQ, FtsI (PBPB), PBPA, and CwsA, CrgA facilitates the recruitment of the proteins essential for peptidoglycan synthesis to the divisome and stabilizes the divisome. CrgA is predicted to have two transmembrane helices. Here, the structure of CrgA was determined in a liquid–crystalline lipid bilayer environment by solid-state NMR spectroscopy. Oriented-sample data yielded orientational restraints, whereas magic-angle spinning data yielded interhelical distance restraints. These data define a complete structure for the transmembrane domain and provide rich information on the conformational ensembles of the partially disordered N-terminal region and interhelical loop. The structure of the transmembrane domain was refined using restrained molecular dynamics simulations in an all-atom representation of the same lipid bilayer environment as in the NMR samples. The two transmembrane helices form a left-handed packing arrangement with a crossing angle of 24° at the conserved Gly39 residue. This helix pair exposes other conserved glycine and alanine residues to the fatty acyl environment, which are potential sites for binding CrgA’s partners such as CwsA and FtsQ. This approach combining oriented-sample and magic-angle spinning NMR spectroscopy in native-like lipid bilayers with restrained molecular dynamics simulations represents a powerful tool for structural characterization of not only isolated membrane proteins, but their complexes, such as those that form macromolecular machines.

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Section: Resultsmentioning

confidence: 96%

Section: Discussionmentioning

confidence: 99%

Section: Significancementioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Structure of CrgA, a cell division structural and regulatory protein fromMycobacterium tuberculosis, in lipid bilayers

Das

Dai

Hung

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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DivIVA concentrates mycobacterial cell envelope assembly for initiation and stabilization of polar growth

et al. 2018

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In many model organisms, diffuse patterning of cell wall peptidoglycan synthesis by the actin homolog MreB enables the bacteria to maintain their characteristic rod shape. In Caulobacter crescentus and Escherichia coli, MreB is also required to sculpt this morphology de novo. Mycobacteria are rod-shaped but expand their cell wall from discrete polar or subpolar zones. In this genus, the tropomyosin-like protein DivIVA is required for the maintenance of cell morphology. DivIVA has also been proposed to direct peptidoglycan synthesis to the tips of the mycobacterial cell. The precise nature of this regulation is unclear, as is its role in creating rod shape from scratch. We find that DivIVA localizes nascent cell wall and covalently associated mycomembrane but is dispensable for the assembly process itself. Mycobacterium smegmatis rendered spherical by peptidoglycan digestion or by DivIVA depletion are able to regain rod shape at the population level in the presence of DivIVA. At the single cell level, there is a close spatiotemporal correlation between DivIVA foci, rod extrusion and concentrated cell wall synthesis. Thus, although the precise mechanistic details differ from other organisms, M. smegmatis also establish and propagate rod shape by cytoskeleton-controlled patterning of peptidoglycan. Our data further support the emerging notion that morphology is a hardwired trait of bacterial cells.

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Intermediate Filaments Supporting Cell Shape and Growth in Bacteria

Kelemen

2017

Subcellular Biochemistry

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For years intermediate filaments (IF), belonging to the third class of filamentous cytoskeletal proteins alongside microtubules and actin filaments, were thought to be exclusive to metazoan cells. Structurally these eukaryote IFs are very well defined, consisting of globular head and tail domains, which flank the central rod-domain. This central domain is dominated by an α-helical secondary structure predisposed to form the characteristic coiled-coil, parallel homo-dimer. These elementary dimers can further associate, both laterally and longitudinally, generating a variety of filament-networks built from filaments in the range of 10 nm in diameter. The general role of these filaments with their characteristic mechano-elastic properties both in the cytoplasm and in the nucleus of eukaryote cells is to provide mechanical strength and a scaffold supporting diverse shapes and cellular functions.Since 2003, after the first bacterial IF-like protein, crescentin was identified, it has been evident that bacteria also employ filamentous networks, other than those built from bacterial tubulin or actin homologues, in order to support their cell shape, growth and, in some cases, division. Intriguingly, compared to their eukaryote counterparts, the group of bacterial IF-like proteins shows much wider structural diversity. The sizes of both the head and tail domains are markedly reduced and there is great variation in the length of the central rod-domain. Furthermore, bacterial rod-domains often lack the sub-domain organisation of eukaryote IFs that is the defining feature of the IF-family. However, the fascinating display of filamentous assemblies, including rope, striated cables and hexagonal laces together with the conditions required for their formation both in vitro and in vivo strongly resemble that of eukaryote IFs suggesting that these bacterial proteins are deservedly classified as part of the IF-family and that the current definition should be relaxed slightly to allow their inclusion. The lack of extensive head and tail domains may well make the bacterial proteins more amenable for structural characterisation, which will be essential for establishing the mechanism for their association into filaments. What is more, the well-developed tools for bacterial manipulations provide an excellent opportunity of studying the bacterial systems with the prospect of making significant progress in our understanding of the general underlying principles of intermediate filament assemblies.

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Characterization of CrgA, a New Partner of the Mycobacterium tuberculosis Peptidoglycan Polymerization Complexes

Cited by 61 publications

References 54 publications

Structure of CrgA, a cell division structural and regulatory protein fromMycobacterium tuberculosis, in lipid bilayers

Structure of CrgA, a cell division structural and regulatory protein fromMycobacterium tuberculosis, in lipid bilayers

DivIVA concentrates mycobacterial cell envelope assembly for initiation and stabilization of polar growth

Intermediate Filaments Supporting Cell Shape and Growth in Bacteria

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