<i><scp>H</scp>aloferax volcanii</i> archaeosortase is required for motility, mating, and <scp>C</scp>‐terminal processing of the <scp>S</scp>‐layer glycoprotein

Halim, Mohd Farid Abdul; Pfeiffer, Friedhelm; Zou, James; Frisch, Andrew; Haft, Daniel H.; Wu, Si; Tolić, Nikola; Brewer, Heather M.; Payne, Samuel H.; Paša‐Tolić, Ljiljana; Pohlschröder, Mechthild

doi:10.1111/mmi.12248

Cited by 59 publications

(68 citation statements)

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“…This glycoprotein was previously thought to be anchored to the cell membrane by the intercalation of a C-terminal TM segment (15). However, we have demonstrated that this TM segment, which is part of a conserved tripartite structure, is processed in an archaeosortase A (ArtA)-dependent manner (14). In light of previous results demonstrating that this extensively studied cell wall subunit is lipid modified with archaetidic acid (2,3-di-O-phytanyl-sn-glycerylphosphate) (16)(17)(18)(19), we hypothesized that ArtA also mediates proteolysis-coupled lipid modification.…”

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confidence: 87%

“…In light of previous results demonstrating that this extensively studied cell wall subunit is lipid modified with archaetidic acid (2,3-di-O-phytanyl-sn-glycerylphosphate) (16)(17)(18)(19), we hypothesized that ArtA also mediates proteolysis-coupled lipid modification. Cells lacking ArtA exhibit a growth defect, are relatively small, and have a severe motility defect (14). While some of these phenotypes are consistent with defects that might be caused by an unstable cell wall due to the improper surface anchoring of the unprocessed SLG, H. volcanii is predicted to contain eight additional ArtA substrates, so the failure to accurately process these other substrates might also contribute to the observed phenotypes (12).…”

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confidence: 94%

“…Protein extraction, LDS-PAGE, and Coomassie blue staining. Protein from cell pellets, CsCl purified samples, and trichloroacetic acid (TCA) precipitations of H. volcanii strains were prepared, separated by lithium dodecyl sulfate-polyacrylamide gel electrophoresis (LDS-PAGE), and stained with Coomassie brilliant blue as previously described (14). Briefly, the liquid cultures were grown until the mid-log phase (OD 600 of ϳ0.5).…”

Section: Methodsmentioning

confidence: 99%

“…An isogenic strain, FH37, containing the trpA gene insertion but having an unaltered slg PGF motif, was constructed and used as wild-type control. (14), using the standard polyethylene glycol (PEG) transformation protocol (20). pTA963 is an overexpression vector derived from pBluescript II with the H. volcanii pHV2 replication origin in addition to both pyrE2 (encoding the product with accession number WP_004044590.1) and hdrB (encoding the product with accession number WP_004044748.1) markers.…”

Section: Methodsmentioning

confidence: 99%

“…The first in vivo characterization of an archaeosortase was carried out in the model archaeon Haloferax volcanii (14), which has a cell wall consisting of a single protein, the S-layer glycoprotein (SLG) (15). This glycoprotein was previously thought to be anchored to the cell membrane by the intercalation of a C-terminal TM segment (15).…”

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Permuting the PGF Signature Motif Blocks both Archaeosortase-Dependent C-Terminal Cleavage and Prenyl Lipid Attachment for the Haloferax volcanii S-Layer Glycoprotein

et al. 2016

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For years, the S-layer glycoprotein (SLG), the sole component of many archaeal cell walls, was thought to be anchored to the cell surface by a C-terminal transmembrane segment. Recently, however, we demonstrated that the Haloferax volcanii SLG C terminus is removed by an archaeosortase (ArtA), a novel peptidase. SLG, which was previously shown to be lipid modified, contains a C-terminal tripartite structure, including a highly conserved proline-glycine-phenylalanine (PGF) motif. Here, we demonstrate that ArtA does not process an SLG variant where the PGF motif is replaced with a PFG motif (slg G796F,F797G ). Furthermore, using radiolabeling, we show that SLG lipid modification requires the PGF motif and is ArtA dependent, lending confirmation to the use of a novel C-terminal lipid-mediated protein-anchoring mechanism by prokaryotes. Similar to the case for the ⌬artA strain, the growth, cellular morphology, and cell wall of the slg G796F,F797G strain, in which modifications of additional H. volcanii ArtA substrates should not be altered, are adversely affected, demonstrating the importance of these posttranslational SLG modifications. Our data suggest that ArtA is either directly or indirectly involved in a novel proteolysis-coupled, covalent lipid-mediated anchoring mechanism. Given that archaeosortase homologs are encoded by a broad range of prokaryotes, it is likely that this anchoring mechanism is widely conserved. IMPORTANCEProkaryotic proteins bound to cell surfaces through intercalation, covalent attachment, or protein-protein interactions play critical roles in essential cellular processes. Unfortunately, the molecular mechanisms that anchor proteins to archaeal cell surfaces remain poorly characterized. Here, using the archaeon H. volcanii as a model system, we report the first in vivo studies of a novel protein-anchoring pathway involving lipid modification of a peptidase-processed C terminus. Our findings not only yield important insights into poorly understood aspects of archaeal biology but also have important implications for key bacterial species, including those of the human microbiome. Additionally, insights may facilitate industrial applications, given that photosynthetic cyanobacteria encode uncharacterized homologs of this evolutionarily conserved enzyme, or may spur development of unique drug delivery systems.A variety of protein complexes decorate cell surfaces, where they support cell stability and facilitate interactions between the cell and the extracellular environment (1-3). Determining the mechanisms that allow these proteins to remain associated with the cell envelope is critical for developing a comprehensive understanding of the biosynthesis of cell surface structures. Protein anchoring to the cell membrane was likely first accomplished by means of transmembrane (TM) segments (4). However, as cell membranes became densely packed with protein complexes, protein-anchoring mechanisms that allowed more economic use of membrane surface area emerged (5). One such mechanism anchors...

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confidence: 87%

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confidence: 94%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

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Permuting the PGF Signature Motif Blocks both Archaeosortase-Dependent C-Terminal Cleavage and Prenyl Lipid Attachment for the Haloferax volcanii S-Layer Glycoprotein

et al. 2016

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S ‐Layers

Engelhardt

2016

Encyclopedia of Life Sciences

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Microbial surface layer (S‐layer) proteins assemble into two‐dimensional (2D) crystalline lattices on the cell surface of many species of Archaea and Bacteria and represent the outermost cell wall component, except for the existence of a carbohydrate capsule. Despite their widespread occurrence, the function of S‐layers remained obscure. Analysing the S‐layer structure on the one hand and the interaction of the 2D crystal with the underlying cell membrane on the other, reveals the S‐layers' cell wall function and the mechanism of cell stabilisation in Archaea . This basic function is not obvious in Bacteria . Here, experimental data suggest a major role in mediating and controlling interactions of S‐layers with the microbes' environment. The hybrid functional role of S‐layers will be understood more clearly if structural research is combined with the investigation of S‐layer interactions with the adjacent cell envelope components and the complex environmental factors. Key Concepts Microbial S‐layers have a hybrid functional role, that is, mediating cell stability and controlling cell surface properties. The architecture of S‐layers and their close interactions with the cell membrane determine and explain the S‐layers' cell wall function in Archaea . Attracting and repelling surface characteristics of S‐layers in Bacteria mediate the interactions with the environment and contribute to the proliferation and survival of cells. The investigation of S‐layer functions requires the combined analysis of the S‐layer structure and its impact on the cell envelope components as well as of interactions with environmental factors.

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Modifying Post‐Translational Modifications: A Strategy Used by Archaea for Adapting to Changing Environments?

Eichler

2020

BioEssays

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In concert with the selective pressures affecting protein folding and function in the extreme environments in which they can exist, proteins in Archaea have evolved to present permanent molecular adaptations at the amino acid sequence level. Such adaptations may not, however, suffice when Archaea encounter transient changes in their surroundings. Post‐translational modifications offer a rapid and reversible layer of adaptation for proteins to cope with such situations. Here, it is proposed that Archaea further augment their ability to survive changing growth conditions by modifying the extent, position, and, where relevant, the composition of different post‐translational modifications, as a function of the environment. Support for this hypothesis comes from recent reports describing how patterns of protein glycosylation, methylation, and other post‐translational modifications of archaeal proteins are altered in response to environmental change. Indeed, adjusting post‐translational modifications as a means to cope with environmental variability may also hold true beyond the Archaea.

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Haloferax volcanii archaeosortase is required for motility, mating, and C‐terminal processing of the S‐layer glycoprotein

Cited by 59 publications

References 39 publications

Permuting the PGF Signature Motif Blocks both Archaeosortase-Dependent C-Terminal Cleavage and Prenyl Lipid Attachment for the Haloferax volcanii S-Layer Glycoprotein

Permuting the PGF Signature Motif Blocks both Archaeosortase-Dependent C-Terminal Cleavage and Prenyl Lipid Attachment for the Haloferax volcanii S-Layer Glycoprotein

S ‐Layers

Modifying Post‐Translational Modifications: A Strategy Used by Archaea for Adapting to Changing Environments?

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