Identification of a collagen type I adhesin of Bacteroides fragilis

Bruna P. G. V. Galvão, Brandon W. Weber, Mohamed S. Rafudeen, Eliane O. Ferreira, Sheila Patrick, Valerie R. Abratt

Research output: Contribution to journalArticle

7 Citations (Scopus)
219 Downloads (Pure)

Abstract

Bacteroides fragilis is an opportunistic pathogen which can cause life threatening infections in humans and animals. The ability to adhere to components of the extracellular matrix, including collagen, is related to bacterial host colonisation. Collagen Far Western analysis of the B. fragilis outer membrane protein (OMP) fraction revealed the presence two collagen adhesin bands of ∼31 and ∼34 kDa. The collagen adhesins in the OMP fraction were separated and isolated by two-dimensional SDS-PAGE and also purified by collagen affinity chromatography. The collagen binding proteins isolated by both these independent methods were subjected to tandem mass spectroscopy for peptide identification and matched to a single hypothetical protein encoded by B. fragilis NCTC 9343 (BF0586), conserved in YCH46 (BF0662) and 638R (BF0633) and which is designated in this study as cbp1 (collagen binding protein). Functionality of the protein was confirmed by targeted insertional mutagenesis of the cbp1 gene in B. fragilis GSH18 which resulted in the specific loss of both the ∼31 kDa and the ∼34 kDa adhesin bands. Purified his-tagged Cbp1, expressed in a B. fragilis wild-type and a glycosylation deficient mutant, confirmed that the cbp1 gene encoded the observed collagen adhesin, and showed that the 34 kDa band represents a glycosylated version of the ∼31 kDa protein. Glycosylation did not appear to be required for binding collagen. This study is the first to report the presence of collagen type I adhesin proteins in B. fragilis and to functionally identify a gene encoding a collagen binding protein.
Original languageEnglish
Article numbere91141
JournalPLoS ONE
Volume9
Issue number3
DOIs
Publication statusPublished - 11 Mar 2014

Keywords

  • adhesin
  • collagen type 1 adhesin
  • unclassified drug
  • affinity chromatolgraphy
  • bacterial gene
  • bacterial outer membrane
  • bacterial strain
  • Bacteroides fragilis
  • cbp1 gene
  • cell count
  • controlled study
  • nonhuman
  • protein analysis
  • protein expression
  • protein glycosylation
  • tandem mass spectromety
  • wild type

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