Identification and characterization of a cell surface protein of Prevotella intermedia 17 with broad-spectrum binding activity for extracellular matrix proteins.
Prevotella intermedia binds and invades a variety of host cells. This binding is most probably mediated through cell surface proteins termed adhesins. To identify proteins binding to the host extracellular matrix (ECM) component, fibronectin, and study the molecular mechanism underlying bacterial colonization, we applied proteomic approaches to perform a global investigation of P. intermedia strain 17 outer membrane proteins. 2-DE followed by Far Western Blot analysis using fibronectin as a probe revealed a 29-kDa fibronectin-binding protein, designated here AdpB. The molecular identity of the protein was determined using PMF followed by a search of the P. intermedia 17 protein database. Database searches revealed the similarity of AdpB to multiple bacterial outer membrane proteins including the fibronectin-binding protein from Campylobacter jejuni. A recombinant AdpB protein bound fibronectin as well as other host ECM components, including fibrinogen and laminin, in a saturable, dose-dependent manner. Binding of AdpB to immobilized fibronectin was also inhibited by soluble fibronectin, laminin, and fibrinogen, indicating the binding was specific. Finally, immunoelectron microscopy with anti-AdpB demonstrated the cell surface location of the protein. This is the first cell surface protein with a broad-spectrum ECM-binding abilities identified and characterized in P. intermedia 17.
Yu
Fan
F
Philips Institute of Oral and Craniofacial Molecular Biology, Virginia Commonwealth University, Richmond, VA 23298-0566, USA.
Iyer
Divya
D
Anaya
Cecilia
C
Lewis
Janina P
JP
D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000378:metabolism*; D003164:Competitive Bidding; D015180:Electrophoresis, Gel, Two-Dimensional; D016326:Extracellular Matrix Proteins / Q000378:metabolism*; D005353:Fibronectins / Q000378:metabolism*; D008969:Molecular Sequence Data; D018713:Prevotella intermedia / Q000378:metabolism*; D011485:Protein Binding; D017386:Sequence Homology, Amino Acid; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods
D001425:Bacterial Outer Membrane Proteins; D016326:Extracellular Matrix Proteins; D005353:Fibronectins