Functional Characterization of the Interaction between Bacterial Adhesin Multivalent Adhesion Molecule 7 (MAM7) Protein and Its Host Cell Ligands

Anne Marie Krachler,Kim Orth

doi:10.1074/jbc.m111.291377

Abstract

The ability of a pathogen to rapidly form a stable interaction with the host cell surface is key to its success. Bacterial pathogens use a repertoire of virulence factors, but their efficient use relies on close contact between the host and the pathogen. We have recently identified a constitutively expressed MAM7 (multivalent adhesion molecule 7), which is widely distributed in gram-negative pathogens and enables them to establish initial contact with the host cell. Here, we describe the dissection of the MAM7 interaction with the host cell surface into two distinct binding events, involving the host protein fibronectin and the membrane phospholipid phosphatidic acid. We analyzed which domains within MAM7 and fibronectin are necessary for complex formation. We further studied phosphatidic acid binding by MAM7 using site-directed mutagenesis and liposome association assays and demonstrated that a specific distribution of basic charge on MAM7 is required for high affinity binding. Finally, we showed that fibronectin and phosphatidic acid binding to MAM7 are not mutually exclusive and that the three molecules likely assemble into a tripartite complex on the host cell surface.

Highlights

The outer membrane multivalent adhesion molecule 7 (MAM7) initiates host cell killing by Gram-negative pathogens
We have demonstrated that MAM7 expressed on the surface of non-pathogenic bacteria may be utilized as a potent inhibitor of pathogen infection in tissue culture
MAM7 is capable of binding both fibronectin and phosphatidic acid on the host cell surface, and both interactions are required for bacterial adhesion to host cells [5]

Summary

Background

The outer membrane multivalent adhesion molecule 7 (MAM7) initiates host cell killing by Gram-negative pathogens. We have recently identified a constitutively expressed MAM7 (multivalent adhesion molecule 7), which is widely distributed in Gram-negative pathogens and enables them to establish initial contact with the host cell. We describe the dissection of the MAM7 interaction with the host cell surface into two distinct binding events, involving the host protein fibronectin and the membrane phospholipid phosphatidic acid. MAM7 is constitutively expressed, enabling Gram-negative pathogens to establish immediate contact with host cells upon their first encounter, which in turn can lead to up-regulation of other, pathogen- and host cell-specific adhesins and virulence factors (5, 8 –10). We studied whether MAM7 is capable of binding to fibronectin and phosphatidic acid simultaneously and found that the three molecules likely form a tripartite complex

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION