Abstract
Listeria monocytogenes causes listeriosis, a potentially fatal food-borne disease. The condition is especially harmful to pregnant women. Listeria outbreaks can originate from diverse foods, highlighting the need for novel strategies to improve food safety. The first step in Listeria invasion is internalization of the bacteria, which is mediated by the interaction of the internalin family of virulence factors with host cell receptors. A crucial interaction for Listeria invasion of the placenta, and thus a target for therapeutic intervention, is between internalin B (InlB) and the receptor c-Met. Single-domain antibodies (VHH, also called nanobodies, or sdAbs) from camel heavy-chain antibodies are a novel solution for preventing Listeria infections. The VHH R303, R330, and R326 all bind InlB with high affinity; however, the molecular mechanism behind their mode of action was unknown. We demonstrate that despite a high degree of sequence and structural diversity, the VHH bind a single epitope on InlB. A combination of gentamicin protection assays and florescent microscopy establish that InlB-specific VHH inhibit Listeria invasion of HeLa cells. A high-resolution X-ray structure of VHH R303 in complex with InlB showed that the VHH binds at the c-Met interaction site on InlB, thereby acting as a competitive inhibitor preventing bacterial invasion. These results point to the potential of VHH as a novel class of therapeutics for the prevention of listeriosis.
Highlights
Listeria monocytogenes causes listeriosis, a potentially fatal food-borne disease
The invasion of nonphagocytic cells by L. monocytogenes occurs through the action of a complex set of virulence factors that allow the bacteria to enter host cells, escape the vacuole, and hijack the actin network to spread from cell to cell [5]
Listeria host cell entry is the initial step in pathogenesis, and it is mediated by two members of the internalin family of virulence factors (InlA2 and internalin B (InlB)) [6, 7]
Summary
Ing pregnancy is further compounded by the capacity of the bacteria to cross the placental barrier, which can result in termination of the developing fetus [4]. Listeria host cell entry is the initial step in pathogenesis, and it is mediated by two members of the internalin family of virulence factors (InlA2 and InlB) [6, 7]. Binding of InlA and InlB to host cell receptors activates signaling cascades that trigger receptormediated endocytosis and internalization of the bacteria. InlA and InlB have different receptors and are responsible for mediating entry into different cell types and biological barriers. The interaction of InlA with the host receptor E-cadherin is important for Listeria penetration of the intestinal barrier and invasion of several epithelial cell types [8]. C-Met functions as the receptor for the hepatocyte growth factor and is required for normal embryonic development, pointing to the importance of InlB in pregnancy-related listeriosis.
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