Effect of Ergosterol on the Binding of KMP-11 with Phospholipid Membranes: Implications in Leishmaniasis

Abstract

Kinetoplastid membrane protein-11 (KMP-11) is a small noble immunogenic protein present in the surfaces of all kinetoplastid protozoans. The parasite membrane comprises ergosterol as the major component. Although the interaction of KMP-11 with membrane and the role of ergosterol have been implicated in the process of infection and in the survival mechanism of the parasite, no molecular level understanding is available. In this paper, we have investigated the binding of KMP-11 with phospholipid model membrane composed of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) in the absence and presence of different molar percentages of ergosterol at different pH. Our results suggest that, at pH 7.5, the binding affinity of WT KMP-11 increased significantly with the increase in the percentage of ergosterol whereas there occurred a large drop of binding at low pH. In order to obtain detailed insights into the conformational positioning of the protein at the membrane in the presence of ergosterol, four different site specific single tryptophan mutants (Y5W, Y48W, F62W, and Y89W) have been generated in which the inserted tryptophan residues (the donor) and dehydroergosterol (DHE, the acceptor)of the membrane constitute FRET pairs. Energy transfer efficiency and donor-acceptor distance calculations suggested strong attachment of the immunogenic protein on the membrane surface. The presented results emphasize that an interconnection between membrane interaction, the presence of ergosterol, and other environmental factors may play crucial roles to ensure the survival of Leishmania parasites inside the host environment.