Biophysical Society Thematic Meeting

Biophysics in the Understanding, Diagnosis, and Treatment of Infectious Diseases Poster Abstracts

45-POS

Board 45

Identification and Characterization of Protein Components of Bacteroides Fragilis

Fimbriae

Bruna Galvao,

Brandon Weber

, Valerie Abratt.

University of Cape Town, Cape Town, South Africa.

Bacterial surface structures involved in attachment, such as fimbriae, are considered important

virulence factors. The aim of this study is to isolate and identify the protein components of the

fimbriae of

Bacteroides fragilis

for structural characterisation. Evaluation of fimbriae production

was performed by growing

B. fragilis

strains on liquid and solid brain heart infusion (BHI) and

Wilkins Chalgren media. Cells were visualised by transmission electron microscopy to assess

which conditions gave rise to the production of fimbriae. Fimbriae were isolated by gentle

shearing from the surface of the bacteria and assessed by TEM and SDS-PAGE. Protein bands of

interest were excised and analysed by LC MS/MS. Antibodies were raised to the recombinant

version of one of the candidate fimbrial proteins and used for immunogold localization studies to

confirm the role of this protein as a component of the observed fimbrial structures.

Production of fimbriae by

B. fragilis

was shown to be dependent on the bacterial strain and

growth conditions used, with the optimum combination observed for strain

B. fragilis

638R

grown on solid BHI medium. These fimbriae were observed by TEM

in situ

on the cells, in the

sheared and precipitated protein fractions. The isolated fimbrial proteins, examined using SDS-

PAGE, revealed four distinct protein bands. These were analysed by LC MS/MS and revealed a

number of candidate fimbrial proteins. One of these, encoded by ORF BF638R_2242, shows

homology to both the FimA and Mfa1 fimbrial proteins of the anaerobic oral

pathogen

Porphyromonas gingivalis

and was chosen for further study. The role of the putative

fimbrial protein as a component of the observed fimbrial structures was investigated by

immunogold labelling studies.

One strong fimbrial gene candidate (FimA) has been identified so far and is the subject of

ongoing research.