title: Investigation of the mechanism of binding between internalin B and heparin using surface plasmon resonance.
abstract: Listeria monocytogenes, a food-borne pathogen that infects immunocompromised patients, enters and proliferates within mammalian cells by taking advantage of host cell machinery. While entry into macrophages and other phagocytic cells occurs constitutively, intracellular invasion of nonphagocytic cells, such as epithelial and endothelial cells, occurs through induced phagocytosis. Invasion of these nonphagocytic cell types is under the control of the secreted L. monocytogenes protein internalin B (InlB), which directly associates with and activates the receptor tyrosine kinase Met. Activation of Met by InlB has previously been shown to be potentiated by binding of glycosaminoglycans to the GW domains of this protein. We studied the interaction between heparin and full-length InlB as well as a truncated, functional form of InlB to understand the mode of interaction between these two molecules. InlB preferred long-chain (>or=dp14) heparin oligosaccharides, and the interaction with heparin fit a complicated binding model with a dissociation constant in the nanomolar range. While there are various explanations for this complicated binding model, one supported by our data involves binding and rebinding of InlB to multiple binding sites on heparin in a positive and weakly cooperative manner. This mode is consistent with enhancement of interaction of InlB with glycosaminoglycans for activation of Met.
authors.affiliation: Department of Chemical and Biochemical Engineering, University of Iowa, Iowa City, Iowa 52242, USA.
authors.forename: Sybil C Lang
authors.initials: SC
authors.lastname: Hrtska
authors.forename: Melissa M
authors.initials: MM
authors.lastname: Kemp
authors.forename: Eva M
authors.initials: EM
authors.lastname: Muñoz
authors.forename: Omaira
authors.initials: O
authors.lastname: Azizad
authors.forename: Mani
authors.initials: M
authors.lastname: Banerjee
authors.forename: Catarina
authors.initials: C
authors.lastname: Raposo
authors.forename: Jyothi
authors.initials: J
authors.lastname: Kumaran
authors.forename: Partho
authors.initials: P
authors.lastname: Ghosh
authors.forename: Robert J
authors.initials: RJ
authors.lastname: Linhardt
mesh_terms: D001426:Bacterial Proteins / Q000378:metabolism*; D001665:Binding Sites; D001667:Binding, Competitive / Q000502:physiology*; D006493:Heparin / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008565:Membrane Proteins / Q000378:metabolism*; D011485:Protein Binding / Q000502:physiology*; D020349:Surface Plasmon Resonance / Q000379:methods*
chemical_list: D001426:Bacterial Proteins; D008565:Membrane Proteins; C095768:inlB protein, Listeria monocytogenes; D006493:Heparin