Listeria monocytogenes adhesion to intestinal cells is mediated, in part, by interaction of Listeria adhesion protein with a eukaryotic cell receptor, heat shock protein 60
Abstract
The 104-kDa Listeria adhesion protein (LAP) in Listeria monocytogenes is involved in binding to various mammalian cell lines. However, the receptor that interacts with LAP in eukaryotic cells is unknown. The goal of this study was identification of the host cell receptor for LAP and its distribution in cells of human intestinal origin. Fluorescent microscopy and scanning immunoelectron microscopy qualitatively demonstrated greater binding capacity of wild type L. monocytogenes strain (F4244) than a LAP-deficient mutant strain (KB208) to Caco-2 cells. Using a Western blot ligand overlay assay, a protein of 58-kDa was identified to be the putative receptor for LAP from Caco-2 cells. N-terminal sequencing and subsequent database search identified this protein as heat shock protein 60 (Hsp60). A modified immunoseparation assay using LAP-specific MAb-H7 and sequential incubation with LAP preparation and Caco-2 lysate confirmed the receptor to be the same 58-kDa protein. Western blot analysis with anti-Hsp60 monoclonal antibody of whole-cell adhesion between Caco-2 and WT also revealed the receptor protein to be a 58-kDa protein, thus corroborating the identification of Hsp60 as a host cell receptor for LAP. Furthermore, the anti-Hsp60 antibody also caused significant 24 to 56% reductions in binding of L. monocytogenes to Caco-2, HT-29, and HCT-8 cells of intestinal origin. Hsp60 is abundantly present in all cell lines tested, however, surface expressed Hsp60 appeared high only in cells of intestinal origin. The adhesion mechanism of L. monocytogenes to eukaryotic cells is a complex process and identification of Hsp60 as a receptor for LAP mainly in cells of intestinal origin suggests the involvement of multiple ligand-receptor interactions to achieve adhesion.
Degree
Ph.D.
Advisors
Bhunia, Purdue University.
Subject Area
Food science|Microbiology|Cellular biology
Off-Campus Purdue Users:
To access this dissertation, please log in to our
proxy server.