<i>Mycobacterium tuberculosis</i>
            FtsX extracellular domain activates the peptidoglycan hydrolase, RipC

Mavrici, Daniela; Marakalala, Mohlopheni J.; Holton, James M.; Prigozhin, Daniil M.; Gee, Christine L.; Zhang, Yanjia J.; Rubín, Eric J.; Alber, Tom

doi:10.1073/pnas.1321812111

Cited by 84 publications

(148 citation statements)

References 29 publications

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“…S1). The structured portions in the N-terminal region (residues [19][20][21][22][23][24][25][26][27] and in the loop (residues 56-67) have remarkably uniform chemical shifts between 70 and 85 ppm and dipolar couplings between 4.3 and 5.5 kHz, with only a couple of outliers for each parameter. The immediate conclusion is these residues form α-helices or β-strands, with the peptide N-H bonds nearly parallel to the bilayer surface, such as an amphipathic helix or a β-sheet in the plane of the bilayer interface.…”

Section: Resultsmentioning

confidence: 99%

“…One is an X-ray structure of the mechanosensitive channel of large conductance, and the other is a single TM helix protein, Rv1761 (19,20). In addition, watersoluble domains of other Mtb membrane proteins have been characterized such as those from PknB and FtsX (21)(22)(23)(24). Although X-ray crystallographers have focused on large membrane proteins, the majority of the 1,162 ORFs of the Mtb genome code for small helical membrane proteins containing one to three TM helices with <40-kDa molecular weight (25).…”

Section: Significancementioning

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: 99%

Section: Significancementioning

confidence: 99%

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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“…FtsEX has been studied in diverse bacterial species for its role in regulating cell wall hydrolysis (9,(44)(45)(46)(47), but its essential role in E. coli cell division, where it was discovered, remained enigmatic because it can be bypassed. Nevertheless, in medium with low osmolarity, FtsEX is essential for division (8,21,31,34).…”

Section: Discussionmentioning

confidence: 99%

“…Although FtsEX is a known component of the divisome in many Gram-negative bacteria, FtsEX in the Gram-positive bacterium Bacillus subtilis is involved in cell wall hydrolysis, but not at the septum (44)(45)(46)(47). Alignment of FtsA sequences from members of both groups revealed a motif (GLTDY) in FtsA of many Gramnegative bacteria that is not present in FtsA of Gram-positive bacteria (Fig.…”

Section: Interaction Between Ftsa and Ftsx Is Important For Divisomementioning

confidence: 98%

FtsEX acts on FtsA to regulate divisome assembly and activity

Pichoff

Lutkenhaus

2016

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

119

203

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Bacterial cell division is driven by the divisome, a ring-shaped protein complex organized by the bacterial tubulin homolog FtsZ. Although most of the division proteins in Escherichia coli have been identified, how they assemble into the divisome and synthesize the septum remains poorly understood. Recent studies suggest that the bacterial actin homolog FtsA plays a critical role in divisome assembly and acts synergistically with the FtsQLB complex to regulate the activity of the divisome. FtsEX, an ATP-binding cassette transporter-like complex, is also necessary for divisome assembly and inhibits division when its ATPase activity is inactivated. However, its role in division is not clear. Here, we find that FtsEX acts on FtsA to regulate both divisome assembly and activity. FtsX interacts with FtsA and this interaction is required for divisome assembly and inhibition of divisome function by ATPase mutants of FtsEX. Our results suggest that FtsEX antagonizes FtsA polymerization to promote divisome assembly and the ATPase mutants of FtsEX block divisome activity by locking FtsA in the inactive form or preventing FtsA from communicating with other divisome proteins. Because FtsEX is known to govern cell wall hydrolysis at the septum, our findings indicate that FtsEX acts on FtsA to promote divisome assembly and to coordinate cell wall synthesis and hydrolysis at the septum. Furthermore, our study provides evidence that FtsA mutants impaired for self-interaction are favored for division, and FtsW plays a critical role in divisome activation in addition to the FtsQLB complex.

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“…RipA has an N-terminal domain that is missing from RipB, but both enzymes are DL-endopeptidases that cleave the bond between the D-glutaminyl and mesodiaminopimelyl residues in the PG peptides (17,18). RipC has a variant Cys-His-His catalytic triad, is activated by FtsX, and has a role in cell wall restructuring during cell division, and a ripC mutant of M. tuberculosis is attenuated in the mouse model (20,21). RipD differs from the other Rip enzymes by having Ala-Ser residues substituted for the canonical Cys-His catalytic residues in the NlpC/P60 domain.…”

mentioning

confidence: 99%

The RipA and RipB Peptidoglycan Endopeptidases Are Individually Nonessential to Mycobacterium smegmatis

Martinelli

Pavelka

2016

J Bacteriol

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Mycobacteria possess a series of Rip peptidoglycan endopeptidases that have been characterized in various levels of detail. The RipA and RipB proteins have been extensively studied and are DL-endopeptidases, and RipA has been considered essential to Mycobacterium smegmatis and Mycobacterium tuberculosis. We show here that the ripA and ripB genes are individually dispensable in M. smegmatis and that at least one of the genes must be expressed for viability. We characterized strains carrying inframe deletion mutations of ripA and ripB and found that both mutant strains exhibited increased susceptibility to a limited number of antibiotics and to detergent but that only the ⌬ripA mutant displayed hypersusceptibility to lysozyme. We also constructed and characterized ⌬ripD and ⌬ripA ⌬ripD mutants and found that the single mutant had only an intermediate lysozyme hypersusceptibility phenotype compared to that of wild-type cells while loss of ripD in the ⌬ripA background partially rescued the antibiotic and lysozyme phenotypes of the ⌬ripA mutant. IMPORTANCEWe show that the RipA endopeptidase, which has been considered essential for cell division in certain mycobacteria, is not essential but that at least it or a similar protein, RipB, must be expressed by the bacteria for viability. This work is the first description of strains carrying single deletion mutations of RipA, RipB, and a novel endopeptidase-like protein, RipD. O ne of the most important characteristics of mycobacteria is a complex cell envelope, the biosynthesis of which is the target of several antimycobacterial drugs (1-3). Our research is focused upon the assembly and maintenance of the peptidoglycan (PG) layer, which surrounds the entirety of the cell and covalently anchors the other components of the cell envelope. The biosynthesis of the PG layer is inhibited by ␤-lactam antibiotics, which have recently been reevaluated as potential antitubercular drugs (4, 5).The mycobacterial PG is composed of glycan strands of Nacetylglucosamine and N-acetyl/N-glycolylmuramic acid with a high degree of interpeptide cross-links (6-9). Assembly is orchestrated by a variety of transglycosylases and transpeptidases, including the penicillin binding proteins (PBPs) PonA1, PonA2, PbpA, and PbpB, as well as several LD-transpeptidases (reviewed in reference 10). In order to incorporate new precursor material into the preexisting cell wall, the old cell wall material must first be metabolized by lytic transglycosylases, endopeptidases, and amidases (11). Cell wall turnover has been extensively studied in Gram-negative bacteria, but comparatively little is known about the balance of degradation/synthesis of the PG of mycobacteria. Most of the research has focused on the Rpf resuscitation-promoting factors, which are PG glycan hydrolases, and on several PG endopeptidases (11,12).The endopeptidases belong to the NlpC/P60 superfamily (13), with the first NlpC/P60 mycobacterial proteins identified in Mycobacterium marinum and given the names iipA and iipB (invasion and ...

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Mycobacterium tuberculosis FtsX extracellular domain activates the peptidoglycan hydrolase, RipC

Cited by 84 publications

References 29 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

FtsEX acts on FtsA to regulate divisome assembly and activity

The RipA and RipB Peptidoglycan Endopeptidases Are Individually Nonessential to Mycobacterium smegmatis

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