Evolution of Streptococcus pyogenes has maximized the efficiency of the Sortase A cleavage motif for cell wall transpeptidation
Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved..
Trafficking of M-protein (Mprt) from the cytosol of Group A Streptococcus pyogenes (GAS) occurs via Sec translocase membrane channels that associate with Sortase A (SrtA), an enzyme that catalyzes cleavage of Mprt at the proximal C-terminal [-LPST355∗GEAA-] motif and subsequent transpeptidation of the Mprt-containing product to the cell wall (CW). These steps facilitate stable exposure of the N-terminus of Mprt to the extracellular milieu where it interacts with ligands. Previously, we found that inactivation of SrtA in GAS cells eliminated Mprt CW transpeptidation but effected little reduction in its cell surface exposure, indicating that the C-terminus of Mprt retained in the cytoplasmic membrane (CM) extends its N-terminus to the cell surface. Herein, we assessed the effects of mutating the Thr355 residue in the WT SrtA consensus sequence (LPST355∗GEAA-) in a specific Mprt, PAM. In vitro, we found that synthetic peptides with mutations (LPSX355GEAA) in the SrtA cleavage site displayed slower cleavage activities with rSrtA than the WT peptide. Aromatic residues at X had the lowest activities. Nonetheless, PAM/[Y355G] still transpeptidated the CW in vivo. However, when using isolated CMs from srtA-inactivated GAS cells, rapid cleavage of PAM/[LPSY355GEAA] occurred at E357∗ but transpeptidation did not take place. These results show that another CM-resident enzyme nonproductively cleaved PAM/[LPSYGE357∗AA]. However, SrtA associated with the translocon channel in vivo cleaved and transpeptidated PAM/[LPSX355∗GEAA] variants. These CM features allow diverse cleavage site variants to covalently attach to the CW despite the presence of other potent nonproductive CM proteases.
| Medienart: |
E-Artikel |
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| Erscheinungsjahr: |
2022 |
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| Erschienen: |
2022 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:298 |
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| Enthalten in: |
The Journal of biological chemistry - 298(2022), 6 vom: 18. Juni, Seite 101940 |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Readnour, Bradley M [VerfasserIn] |
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| Links: |
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| Anmerkungen: |
Date Completed 29.06.2022 Date Revised 14.02.2024 published: Print-Electronic Citation Status MEDLINE |
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| doi: |
10.1016/j.jbc.2022.101940 |
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| funding: |
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| Förderinstitution / Projekttitel: |
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| PPN (Katalog-ID): |
NLM339596937 |
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| 100 | 1 | |a Readnour, Bradley M |e verfasserin |4 aut | |
| 245 | 1 | 0 | |a Evolution of Streptococcus pyogenes has maximized the efficiency of the Sortase A cleavage motif for cell wall transpeptidation |
| 264 | 1 | |c 2022 | |
| 336 | |a Text |b txt |2 rdacontent | ||
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| 500 | |a Date Completed 29.06.2022 | ||
| 500 | |a Date Revised 14.02.2024 | ||
| 500 | |a published: Print-Electronic | ||
| 500 | |a Citation Status MEDLINE | ||
| 520 | |a Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved. | ||
| 520 | |a Trafficking of M-protein (Mprt) from the cytosol of Group A Streptococcus pyogenes (GAS) occurs via Sec translocase membrane channels that associate with Sortase A (SrtA), an enzyme that catalyzes cleavage of Mprt at the proximal C-terminal [-LPST355∗GEAA-] motif and subsequent transpeptidation of the Mprt-containing product to the cell wall (CW). These steps facilitate stable exposure of the N-terminus of Mprt to the extracellular milieu where it interacts with ligands. Previously, we found that inactivation of SrtA in GAS cells eliminated Mprt CW transpeptidation but effected little reduction in its cell surface exposure, indicating that the C-terminus of Mprt retained in the cytoplasmic membrane (CM) extends its N-terminus to the cell surface. Herein, we assessed the effects of mutating the Thr355 residue in the WT SrtA consensus sequence (LPST355∗GEAA-) in a specific Mprt, PAM. In vitro, we found that synthetic peptides with mutations (LPSX355GEAA) in the SrtA cleavage site displayed slower cleavage activities with rSrtA than the WT peptide. Aromatic residues at X had the lowest activities. Nonetheless, PAM/[Y355G] still transpeptidated the CW in vivo. However, when using isolated CMs from srtA-inactivated GAS cells, rapid cleavage of PAM/[LPSY355GEAA] occurred at E357∗ but transpeptidation did not take place. These results show that another CM-resident enzyme nonproductively cleaved PAM/[LPSYGE357∗AA]. However, SrtA associated with the translocon channel in vivo cleaved and transpeptidated PAM/[LPSX355∗GEAA] variants. These CM features allow diverse cleavage site variants to covalently attach to the CW despite the presence of other potent nonproductive CM proteases | ||
| 650 | 4 | |a Journal Article | |
| 650 | 4 | |a Research Support, N.I.H., Extramural | |
| 650 | 4 | |a Streptococcus pyogenes | |
| 650 | 4 | |a bacteria | |
| 650 | 4 | |a bacterial pathogenesis | |
| 650 | 4 | |a c2ll membrane enzymes | |
| 650 | 4 | |a cell surface | |
| 650 | 4 | |a cell wall | |
| 650 | 4 | |a ligand-binding protein | |
| 650 | 4 | |a mutagenesis | |
| 650 | 4 | |a plasminogen | |
| 650 | 4 | |a subcellular fractionation | |
| 650 | 7 | |a Bacterial Proteins |2 NLM | |
| 650 | 7 | |a Aminoacyltransferases |2 NLM | |
| 650 | 7 | |a EC 2.3.2.- |2 NLM | |
| 650 | 7 | |a sortase A |2 NLM | |
| 650 | 7 | |a EC 2.3.2.- |2 NLM | |
| 650 | 7 | |a Cysteine Endopeptidases |2 NLM | |
| 650 | 7 | |a EC 3.4.22.- |2 NLM | |
| 700 | 1 | |a Ayinuola, Yetunde A |e verfasserin |4 aut | |
| 700 | 1 | |a Russo, Brady T |e verfasserin |4 aut | |
| 700 | 1 | |a Liang, Zhong |e verfasserin |4 aut | |
| 700 | 1 | |a Lee, Shaun W |e verfasserin |4 aut | |
| 700 | 1 | |a Ploplis, Victoria A |e verfasserin |4 aut | |
| 700 | 1 | |a Fischetti, Vincent A |e verfasserin |4 aut | |
| 700 | 1 | |a Castellino, Francis J |e verfasserin |4 aut | |
| 773 | 0 | 8 | |i Enthalten in |t The Journal of biological chemistry |d 1945 |g 298(2022), 6 vom: 18. Juni, Seite 101940 |w (DE-627)NLM000004995 |x 1083-351X |7 nnns |
| 773 | 1 | 8 | |g volume:298 |g year:2022 |g number:6 |g day:18 |g month:06 |g pages:101940 |
| 856 | 4 | 0 | |u http://dx.doi.org/10.1016/j.jbc.2022.101940 |3 Volltext |
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