Structural basis of cell wall anchoring by SLH domains in Paenibacillus alvei

Blackler, R.J.; López-Guzmán, Arturo; Hager, Fiona F; Janesch, Bettina; Martinz, Gudrun; Gagnon, S.; Haji-Ghassemi, O.; Kosma, Paul; Messner, Paul; Schäffer, Christina; Evans, Stephen V.

doi:10.1038/s41467-018-05471-3

Cited by 34 publications

(70 citation statements)

References 61 publications

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“…The C‐terminal region is unstructured and solvent exposed on the outward facing surface (Baranova et al, 2012), and it has been used previously as a site for embellishment (Moll et al, 2002). The N‐terminal region of SbsB contains the Surface Layer Homology domain, which is involved in anchoring the S‐layer to the cell‐wall (Blackler et al, 2018). The SbsB crystal structure lacks electron density for the N‐terminal region.…”

Section: Resultsmentioning

confidence: 99%

Programmable assembly of 2D crystalline protein arrays into covalently stacked 3D bionanomaterials

Manea

Garda

Rad

et al. 2020

Biotech & Bioengineering

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Rational embellishment of self‐assembling two‐dimensional (2D) proteins make it possible to build 3D nanomaterials with novel catalytic, optoelectronic and mechanical properties. However, introducing multiple sites of embellishment into 2D protein arrays without affecting the self‐assembly is challenging, limiting the ability to program in additional functionality and new 3D configurations. Here we introduce two orthogonal covalent linkages at multiple sites in a 2D crystalline‐forming protein without affecting its self‐assembly. We first engineered the surface‐layer protein SbsB from Geobacillus stearothermophilus pV72/p2 to display isopeptide bond‐forming protein conjugation pairs, SpyTag or SnoopTag, at four positions spaced 5.7‐10.5 nm apart laterally and 3 nm axially. The C‐terminal and two newly‐identified locations within SbsB monomer accommodated the short SpyTag or SnoopTag peptide tags without affecting the 2D lattice structure. Introducing tags at distinct locations enabled orthogonal and covalent binding of SpyCatcher‐ or SnoopCatcher‐protein fusions to micron‐sized 2D nanosheets. By introducing different types of bifunctional cross‐linkers, the dual‐functionalized nanosheets were programmed to self‐assemble into different 3D stacks, all of which retain their nanoscale order. Thus, our work creates a modular protein platform that is easy to program to create dual‐functionalized 2D and lamellar 3D nanomaterials with new catalytic, optoelectronic, and mechanical properties.

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Section: Resultsmentioning

confidence: 99%

Programmable assembly of 2D crystalline protein arrays into covalently stacked 3D bionanomaterials

Manea

Garda

Rad

et al. 2020

Biotech & Bioengineering

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“…Importantly, pyruvylation imparts an anionic character to the glycoconjugates, which is pivotal to many biological functions. Described functions include the influence on the viscosity of the EPS, bacterial symbiosis with plants [18,28,46], immunostimulatory effects (mostly of CPSs [7,93]), employment of sialylation-like properties in human-type oligosaccharides [198], and cell wall anchoring relying on the Pyr-β- d -ManNAc epitope [14,99,104,195], to name a few. However, learning more about the biological significance of pyruvylated glycoconjugates and delineating a possible association between the position of pyruvylation and functionality are remaining challenges for future research.…”

Section: Discussionmentioning

confidence: 99%

“…These SCWPs are 5–20 kDa in size, composed of species–specific repeats, but lack repetitive alditol phosphates and phosphodiester bonds typical of WTAs and LTAs [100,101,102]—hence the terminology “non-classical” SCWPs. Importantly, they contain a 4,6-ketal pyruvylated β- d -ManNAc residue (4,6Pyr-β- d -ManNAc), imparting a negative charge and serving as a specific cell wall ligand for S-layer homology (SLH) domains usually present in triplicate at the termini of cell surface proteins [99,103,104]. Among such proteins are S-layer proteins, which self-assemble into 2D crystalline arrays on the bacterial cell surface [105,106]; they are important for many biological functions such as maintenance of cell integrity, enzyme display, protection to phagocytosis, and interactions with the host and its immune system [107].…”

Section: Ketal-pyruvylated Glycoconjugatesmentioning

confidence: 99%

“…P. alvei CsaB catalyses the pyruvate modification on a β- d -ManNAc residue present in every SCWP repeat; the resulting 4,6-β- d -ManNAc is the essential epitope for the binding of the bacterium’s SLH domain-containing S-layer protein SpaA, as revealed from the co-crystal structure of synthetic pyruvylated ligand with truncated SpaA SLH [104,194]. Supporting data comes from isothermal titration calorimetry, revealing binding between these modules only when the pyruvate entity was present [104].…”

Section: Ketal-pyruvyltransferasesmentioning

confidence: 99%

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Pyruvate Substitutions on Glycoconjugates

Hager

Sützl

Stefanović

et al. 2019

IJMS

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Glycoconjugates are the most diverse biomolecules of life. Mostly located at the cell surface, they translate into cell-specific “barcodes” and offer a vast repertoire of functions, including support of cellular physiology, lifestyle, and pathogenicity. Functions can be fine-tuned by non-carbohydrate modifications on the constituting monosaccharides. Among these modifications is pyruvylation, which is present either in enol or ketal form. The most commonly best-understood example of pyruvylation is enol-pyruvylation of N-acetylglucosamine, which occurs at an early stage in the biosynthesis of the bacterial cell wall component peptidoglycan. Ketal-pyruvylation, in contrast, is present in diverse classes of glycoconjugates, from bacteria to algae to yeast—but not in humans. Mild purification strategies preventing the loss of the acid-labile ketal-pyruvyl group have led to a collection of elucidated pyruvylated glycan structures. However, knowledge of involved pyruvyltransferases creating a ring structure on various monosaccharides is scarce, mainly due to the lack of knowledge of fingerprint motifs of these enzymes and the unavailability of genome sequences of the organisms undergoing pyruvylation. This review compiles the current information on the widespread but under-investigated ketal-pyruvylation of monosaccharides, starting with different classes of pyruvylated glycoconjugates and associated functions, leading to pyruvyltransferases, their specificity and sequence space, and insight into pyruvate analytics.

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“…The SbsB S-layer protein of Geobacillus stearothermophilus PV72p2 (PDB accession number 4aq1) (Baranova et al 2012) and cell wall-anchoring protein of P. alvei (PDB accession number 6cwc for the SLH domains) (Blackler et al 2018) were used as the templates for the construction of a three-dimensional structural model of P. curdlanolyticus B-6 S1 in the SWISS-MODEL protein-modeling server (http://swissmodel.expasy.org/), representing the unknown and SLH domains of S1, respectively.…”

Section: Methodsmentioning

confidence: 99%

Molecular characterization of hypothetical scaffolding-like protein S1 in multienzyme complex produced by Paenibacillus curdlanolyticus B-6

et al. 2019

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Paenibacillus curdlanolyticus B-6 produces an extracellular multienzyme complex containing a hypothetical scaffolding-like protein and several xylanases and cellulases. The largest (280-kDa) component protein, called S1, has cellulose-binding ability and xylanase activity, thus was considered to function like the scaffolding proteins found in cellulosomes. S1 consists of 863 amino acid residues with predicted molecular mass 91,029 Da and includes two N-terminal surface layer homology (SLH) domains, but most of its sequence shows no homology with proteins of known function. Native S1 (nS1) was highly glycosylated. Purified nS1 and recombinant Xyn11A (rXyn11A) as a major xylanase subunit could assemble in a complex, but recombinant S1 (rS1) could not interact with rXyn11A, indicating that S1 glycosylation is necessary for assembly of the multienzyme complex. nS1 and rS1 showed weak, typical endo-xylanase activity, even though they have no homology with known glycosyl hydrolase family enzymes. S1 and its SLH domains bound tightly to the peptide-glycan layer of P. curdlanolyticus B-6, microcrystalline cellulose, and insoluble xylan, indicating that the SLHs of S1 bind to carbohydrate polymers and the cell surface. When nS1 and rXyn11A were co-incubated with birchwood xylan, the degradation ability was synergistically increased compared with that for each protein; however synergy was not observed for rS1 and rXynA. These results indicate that S1 may have a scaffolding protein-like function by interaction with enzyme subunits and polysaccharides through its glycosylated sites and SLH domains.

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Structural basis of cell wall anchoring by SLH domains in Paenibacillus alvei

Cited by 34 publications

References 61 publications

Programmable assembly of 2D crystalline protein arrays into covalently stacked 3D bionanomaterials

Programmable assembly of 2D crystalline protein arrays into covalently stacked 3D bionanomaterials

Pyruvate Substitutions on Glycoconjugates

Molecular characterization of hypothetical scaffolding-like protein S1 in multienzyme complex produced by Paenibacillus curdlanolyticus B-6

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