Difluorination is an attractive modification to control conformations of hydrocarbon chains. For protein-lipid binding this can have two effects; pre-organisational entropy gain, and creation of additional weak interaction with the protein with minimal change to the desolvation energy. To test the hypothesis that selective fluorination can be used rationally to improve affinity, we study the protein-lipid system CD1d-KRN7000. Presentation of glycolipids by CD1d proteins on the surface of antigen presenting cells triggers potent functional responses from invariant natural killer T (iNKT) cells. Modifying the glycolipid-CD1d interaction through acyl chain modifications can modulate the functional response of iNKT cells. The a-galactosylceramide KRN7000, which is a known CD1d ligand and potent iNKT antigen, has been used in a number of phase I clinical trials in cancer, malaria, tuberculosis, and autoimmune disease. Hence, understanding structure-function relationships of KRN7000 with regard to lipid-protein interaction is a highly relevant problem. A classical molecular dynamics trajectory of the bound lipid was analysed for potential fluorination positions with two purposes: to identify the greatest contribution to pre-organisational entropy gain, and for potential fluorination sites that would have the greatest increase in binding affinity through fluorine-protein interactions. MD simulations were performed on these difluorinated structures and large-scale density functional theory calculations were used to analyse the stereo-electronic effects of the fluorination pattern on the conformation, as well as the specific interactions that the fluorines make with the protein. Close collaboration with experimentalists has led to several of the suggested KRN7000 fluorinated derivatives being synthesised and tested.The results of these assays will be discussed in the context of our predictions.

Using fluorine to optimise affinity: Direct interactions and pre-organisation in lipid binding to CD1d

248th National Meeting (2014)

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