SignalP 6.0 predicts all five types of signal peptides using protein language models

Teufel, Felix; Armenteros, José Juan Almagro; Johansen, Alexander Rosenberg; Gíslason, Magnús Halldór; Pihl, Silas Irby; Tsirigos, Konstantinos D.; Winther, Ole; Brunak, Søren; Heijne, Gunnar von; Nielsen, Henrik

doi:10.1038/s41587-021-01156-3

Cited by 1,466 publications

(1,025 citation statements)

References 36 publications

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“…HHpred raw data was parsed using in-house scripts to generate the architecture of each protein in terms of non-overlapping homologous regions to 2J58 and 3P42. To identify the type of signal peptide and the cleavage site location, each OPX protein was subject to SignalP 6.0 analysis (Teufel et al ., 2022) and to predict the membrane topology of the proteins, TMHMM (Server v. 2.0) (Krogh et al ., 2001) was used.…”

Section: Methodsmentioning

confidence: 99%

Bacterial outer-membrane polysaccharide export (OPX) proteins occupy three structural classes with selective β-barrel porin requirements for polymer secretion

Saïdi

Mahanta

Panda

et al. 2022

Preprint

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Secretion of high-molecular-weight polysaccharides across the bacterial envelope is ubiquitous as it enhances prokaryotic survival in (a)biotic settings. Such polymers are often assembled by Wzx/Wzy- or ABC transporter-dependent schemes that implicate outer-membrane (OM) polysaccharide export (OPX) proteins in polymer translocation to the cell surface. In the social predatory bacterium Myxococcus xanthus, exopolysaccharide (EPS)-pathway WzaX, major spore coat (MASC)-pathway WzaS, and biosurfactant polysaccharide-pathway WzaB were herein found to be truncated OPX homologues of Escherichia coli Wza lacking OM-spanning α-helices. Comparative genomics across all bacteria, complemented with cryo-electron tomography cell-envelope analyses, revealed WzaX/S/B architecture to be the most common amongst three defined OPX-protein structural classes independent of periplasmic thickness. Fold-recognition and deep-learning analyses revealed the conserved M. xanthus proteins MXAN_7418/3226/1916 (encoded adjacent to WzaX/S/B) to be integral OM β-barrels, with structural homology to the poly-N-acetyl-D-glucosamine synthase-dependent pathway porin PgaA. Such porins were identified in bacteria near numerous genes for all three OPX-protein classes. Interior MXAN_7418/3226/1916 β-barrel electrostatics were found to match known properties of their associated polymers. With MXAN_3226 essential for MASC export, and MXAN_7418 absence shown herein to compromise EPS translocation, these data support a novel secretion paradigm for Wzx/Wzy-dependent pathways in which those containing an OPX component that cannot span the OM instead utilize a β-barrel porin to mediate polysaccharide transport across the OM.

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Section: Methodsmentioning

confidence: 99%

Bacterial outer-membrane polysaccharide export (OPX) proteins occupy three structural classes with selective β-barrel porin requirements for polymer secretion

Saïdi

Mahanta

Panda

et al. 2022

Preprint

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“…Alignment was prepared with Clustal Omega online ( https://www.ebi.ac.uk/Tools/msa/clustalo/ ; accessed January 15, 2022). The following elements are indicated, from N to C end, on SntA precursor sequence: signal peptide (gray; amino acids 1–27) as predicted by SignalP 6.0 ( Teufel et al, 2022 ); metallophos domain (green; amino acids 113–426); interdomain linker (orange; amino acids 427–446); 5_nucleotid_C domain (cyan; amino acids 447–680); and C-terminal sorting signal (gray) comprising a LPXTG motif (amino acids 783–787) and a hydrophobic domain with a positively charged tail (amino acids 792–813). SntA core used to run substrate specificity and kinetics experiments corresponds to amino acids 111–727 (shown within borders), which align well with mature CpdB.…”

Section: Introductionmentioning

confidence: 99%

Enzyme Characterization of Pro-virulent SntA, a Cell Wall-Anchored Protein of Streptococcus suis, With Phosphodiesterase Activity on cyclic-di-AMP at a Level Suited to Limit the Innate Immune System

et al. 2022

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Streptococcus suis and Streptococcus agalactiae evade the innate immune system of the infected host by mechanisms mediated by cell wall-anchored proteins: SntA and CdnP, respectively. The former has been reported to interfere with complement responses, and the latter dampens STING-dependent type-I interferon (IFN) response by hydrolysis of bacterial cyclic-di-AMP (c-di-AMP). Both proteins are homologous but, while CdnP has been studied as a phosphohydrolase, the enzyme activities of SntA have not been investigated. The core structure of SntA was expressed in Escherichia coli as a GST-tagged protein that, after affinity purification, was characterized as phosphohydrolase with a large series of substrates. This included 3′-nucleotides, 2′,3′-cyclic nucleotides, cyclic and linear dinucleotides, and a variety of phosphoanhydride or phosphodiester compounds, most of them previously considered as substrates of E. coli CpdB, a periplasmic protein homologous to SntA and CdnP. Catalytic efficiency was determined for each SntA substrate, either by dividing parameters kcat/KM obtained from saturation curves or directly from initial rates at low substrate concentrations when saturation curves could not be obtained. SntA is concluded to act as phosphohydrolase on two groups of substrates with efficiencies higher or lower than ≈ 105 M–1 s–1 (average value of the enzyme universe). The group with kcat/KM ≥ 105 M–1 s–1 (good substrates) includes 3′-nucleotides, 2′,3′-cyclic nucleotides, and linear and cyclic dinucleotides (notably c-di-AMP). Compounds showing efficiencies <104 M–1 s–1 are considered poor substrates. Compared with CpdB, SntA is more efficient with its good substrates and less efficient with its poor substrates; therefore, the specificity of SntA is more restrictive. The efficiency of the SntA activity on c-di-AMP is comparable with the activity of CdnP that dampens type-I IFN response, suggesting that this virulence mechanism is also functional in S. suis. SntA modeling revealed that Y530 and Y633 form a sandwich with the nitrogen base of nucleotidic ligands in the substrate-binding site. Mutants Y530A-SntA, Y633A-SntA, and Y530A+Y633A-SntA were obtained and kinetically characterized. For orientation toward the catalytic site, one tyrosine is enough, although this may depend on the substrate being attacked. On the other hand, both tyrosines are required for the efficient binding of good SntA substrates.

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“…The signal peptide of the PNX precursors is unusual. We could only detect it by SignalP3.0 ( Bendtsen et al 2004 ) and not by SignalP4.1 ( Petersen et al 2011 ), SignalP5.0 ( Almagro-Armenteros et al 2019b ), or SignalP6.0 ( Teufel et al 2022 ). These newer versions of the software failed to identify the signal peptide even in the human PNX precursor protein where propeptide cleavage has been experimentally demonstrated ( Yosten et al 2013a ) or in the PNX precursor from N. norvegicus ( Nguyen et al 2018 ).…”

Section: Resultsmentioning

confidence: 92%

Premetazoan Origin of Neuropeptide Signaling

Yáñez-Guerra

Thiel

Jékely

2022

Molecular Biology and Evolution

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Neuropeptides are a diverse class of signalling molecules in metazoans. They occur in all animals with a nervous system and also in neuron-less placozoans. However, their origin has remained unclear because no neuropeptide shows deep homology across lineages and none have been found in sponges. Here, we identify two neuropeptide precursors, phoenixin and nesfatin, with broad evolutionary conservation. By database searches, sequence alignments and gene-structure comparisons we show that both precursors are present in bilaterians, cnidarians, ctenophores and sponges. We also found phoenixin and a secreted nesfatin precursor homolog in the choanoflagellate Salpingoeca rosetta. Phoenixin in particular, is highly conserved, including its cleavage sites, suggesting that prohormone processing occurs also in choanoflagellates. In addition, based on phyletic patterns and negative pharmacological assays we question the originally proposed GPR-173 (SREB3) as a phoenixin receptor. Our findings revealed that secreted neuropeptide homologs derived from longer precursors have pre-metazoan origins and thus evolved before neurons.

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SignalP 6.0 predicts all five types of signal peptides using protein language models

Cited by 1,466 publications

References 36 publications

Bacterial outer-membrane polysaccharide export (OPX) proteins occupy three structural classes with selective β-barrel porin requirements for polymer secretion

Bacterial outer-membrane polysaccharide export (OPX) proteins occupy three structural classes with selective β-barrel porin requirements for polymer secretion

Enzyme Characterization of Pro-virulent SntA, a Cell Wall-Anchored Protein of Streptococcus suis, With Phosphodiesterase Activity on cyclic-di-AMP at a Level Suited to Limit the Innate Immune System

Premetazoan Origin of Neuropeptide Signaling

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