PG1058 is a novel multidomain protein component of the bacterial type IX secretion system

Jacqueline E. Heath, Christine A. Seers, Paul D. Veith, Catherine A. Butler, Nor Muhammad Nor Azlan, Yu Yen Chen, Nada Slakeski, Benjamin Peng, Lianyi Zhang, Stuart G. Dashper, Keith J. Cross, Steven M. Cleal, Caroline Moore, Eric C. Reynolds

Research output: Contribution to journalArticle

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Abstract

Porphyromonas gingivalis utilises the Bacteroidetes-specific type IX secretion system (T9SS) to export proteins across the outer membrane (OM), including virulence factors such as the gingipains. The secreted proteins have a conserved carboxy-terminal domain essential for type IX secretion that is cleaved upon export. In P. gingivalis the T9SS substrates undergo glycosylation with anionic lipopolysaccharide (A-LPS) and are attached to the OM. In this study, comparative analyses of 24 Bacteroidetes genomes identified ten putative novel components of the T9SS in P. gingivalis, one of which was PG1058. Computer modelling of the PG1058 structure predicted a novel N- to C-terminal architecture comprising a tetratricopeptide repeat (TPR) domain, a β-propeller domain, a carboxypeptidase regulatory domain-like fold (CRD) and an OmpA-C-like putative peptidoglycan binding domain. Inactivation of pg1058 in P. gingivalis resulted in loss of both colonial pigmentation and surface-associated proteolytic activity; a phenotype common to T9SS mutants. Immunoblot and LC-MS/MS analyses of subcellular fractions revealed T9SS substrates accumulated within the pg1058 mutant periplasm whilst whole-cell ELISA showed the Kgp gingipain was absent from the cell surface, confirming perturbed T9SS function. Immunoblot, TEM and whole-cell ELISA analyses indicated A-LPS was produced and present on the pg1058 mutant cell surface although it was not linked to T9SS substrate proteins. This indicated that PG1058 is crucial for export of T9SS substrates but not for the translocation of A-LPS. PG1058 is a predicted lipoprotein and was localised to the periplasmic side of the OM using whole-cell ELISA, immunoblot and LC-MS/MS analyses of subcellular fractions. The structural prediction and localisation of PG1058 suggests that it may have a role as an essential scaffold linking the periplasmic and OM components of the T9SS.

Original languageEnglish
Article number0164313
JournalPLoS One
Volume11
Issue number10
DOIs
Publication statusPublished - 1 Oct 2016

Fingerprint

Bacterial Secretion Systems
Porphyromonas gingivalis
gingipain
secretion
Membranes
Lipopolysaccharides
lipopolysaccharides
Bacteroidetes
Substrates
subcellular fractions
Subcellular Fractions
Enzyme-Linked Immunosorbent Assay
enzyme-linked immunosorbent assay
Proteins
proteins
mutants
cells
Glycosylation
Carboxypeptidases
Peptidoglycan

ASJC Scopus subject areas

  • Medicine(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)

Cite this

Heath, J. E., Seers, C. A., Veith, P. D., Butler, C. A., Nor Azlan, N. M., Chen, Y. Y., ... Reynolds, E. C. (2016). PG1058 is a novel multidomain protein component of the bacterial type IX secretion system. PLoS One, 11(10), [0164313]. https://doi.org/10.1371/journal.pone.0164313

PG1058 is a novel multidomain protein component of the bacterial type IX secretion system. / Heath, Jacqueline E.; Seers, Christine A.; Veith, Paul D.; Butler, Catherine A.; Nor Azlan, Nor Muhammad; Chen, Yu Yen; Slakeski, Nada; Peng, Benjamin; Zhang, Lianyi; Dashper, Stuart G.; Cross, Keith J.; Cleal, Steven M.; Moore, Caroline; Reynolds, Eric C.

In: PLoS One, Vol. 11, No. 10, 0164313, 01.10.2016.

Research output: Contribution to journalArticle

Heath, JE, Seers, CA, Veith, PD, Butler, CA, Nor Azlan, NM, Chen, YY, Slakeski, N, Peng, B, Zhang, L, Dashper, SG, Cross, KJ, Cleal, SM, Moore, C & Reynolds, EC 2016, 'PG1058 is a novel multidomain protein component of the bacterial type IX secretion system', PLoS One, vol. 11, no. 10, 0164313. https://doi.org/10.1371/journal.pone.0164313
Heath, Jacqueline E. ; Seers, Christine A. ; Veith, Paul D. ; Butler, Catherine A. ; Nor Azlan, Nor Muhammad ; Chen, Yu Yen ; Slakeski, Nada ; Peng, Benjamin ; Zhang, Lianyi ; Dashper, Stuart G. ; Cross, Keith J. ; Cleal, Steven M. ; Moore, Caroline ; Reynolds, Eric C. / PG1058 is a novel multidomain protein component of the bacterial type IX secretion system. In: PLoS One. 2016 ; Vol. 11, No. 10.
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