Fluorine’s outstanding capability to influence physico-chemical properties of organic molecules make fluorine substitution an attractive strategy to modulate the binding affinities in protein-ligand and protein-protein systems. The direction and magnitude with which fluorine influences the binding affinity is often difficult to predict, as multiple effects may play a role. Computational methods and Molecular Dynamics (MD) simu- lations in particular are an excellent tool to study such effects, as they allow for atomic precision and in-depth insights into the multiple components of the binding affinity. I have compiled selected aspects of using computational methods to study fluorination in protein systems in a perspective article, which is part of this thesis. Furthermore, I use MD simulations to study how fluorine impacts the binding properties of selected protein- protein or protein-amino acid systems. The first system is the complex of trypsin with fluorinated variants of Abu-BPTI. In Abu-BPTI, the crucial amino acid Lys15 is replaced by the shorter aliphatic amino acid Abu, which comes with a loss of inhibitor strength. Some of the inhibitor strength can be restored by fluorination. An MD based study of the water molecules in the binding pocket of these protein complexes revealed a highly dynamic water network, with strongly interconnected water molecules. The fluorinated moiety of Abu does not interact with the water molecules, making it unlikely that the in- hibitor activity is restored via water mediated bonds between trypsin and fluorine. The analysis of the unbinding paths using Random Acceleration MD (RAMD) revealed a novel metastable state for all of the studied variants, which we call the pre-bound state. This state is clearly distinct from the fully bound state in position and orientation and is stable in long MD simulations. Moreover, the states differ in their interaction pattern, with fluorine possibly having a stabilizing effect on the formation of the pre-bound state. The second system studied here is the protein PTP1B in complex with a phosphoty- rosine mimetic with a highly fluorinated headgroup. I characterized the binding pose and fluorine specific interactions between the mimetic and the protein binding pocket residues. The third system is the GrsA A domain in complex with fluorinated pheny- lalanine variants. Here, I characterized the interruption of an aromatic interaction by fluorination. With respect to the impact of fluorination on the interaction of proteins, the results of this thesis show no evidence for direct water mediated interactions between proteins and fluorine, but indicate that fluorine might influence pre-bound intermedi- ates. For future research it might be interesting to investigate the effects on other components of binding affinities, such as binding entropy. Furthermore, not only the impact of fluorination on fully bound states should be investigated, but also the impact on possible pre-bound states.
