Abstract
Biomolecular recognition between proteins follows complex mechanisms, the understanding of which can substantially advance drug discovery efforts. Here, we track each step of the binding process in atomistic detail with molecular dynamics simulations using trypsin and its inhibitor bovine pancreatic trypsin inhibitor (BPTI) as a model system. We use umbrella sampling to cover a range of unbinding pathways. Starting from these simulations, we subsequently seed classical simulations at different stages of the process and combine them to a Markov state model. We clearly identify three kinetically separated states (an unbound state, an encounter state, and the final complex) and describe the mechanisms that dominate the binding process. From our model, we propose the following sequence of events. The initial formation of the encounter complex is driven by long-range interactions because opposite charges in trypsin and BPTI draw them together. The encounter complex features the prealigned binding partners with binding sites still partially surrounded by solvation shells. Further approaching leads to desolvation and increases the importance of van der Waals interactions. The native binding pose is adopted by maximizing short-range interactions. Thereby side-chain rearrangements ensure optimal shape complementarity. In particular, BPTI’s P1 residue adapts to the S1 pocket and prime site residues reorient to optimize interactions. After the paradigm of conformation selection, binding-competent conformations of BPTI and trypsin are already present in the apo ensembles and their probabilities increase during this proposed two-step association process. This detailed characterization of the molecular forces driving the binding process includes numerous aspects that have been discussed as central to the binding of trypsin and BPTI and protein complex formation in general. In this study, we combine all these aspects into one comprehensive model of protein recognition. We thereby contribute to enhance our general understanding of this fundamental mechanism, which is particularly critical as the development of biopharmaceuticals continuously gains significance.
Highlights
The growing relevance of biopharmaceuticals [1] renders a comprehensive understanding of the fundamental mechanisms of protein-protein association, recognition, and binding of utmost importance
We investigate the binding process between trypsin and its inhibitor bovine pancreatic trypsin inhibitor (BPTI) and aim to understand the fundamental factors that contribute to recognition and binding in atomic detail
We suggest a two-step binding mechanism following the paradigm of conformational selection with a population shift
Summary
The growing relevance of biopharmaceuticals [1] renders a comprehensive understanding of the fundamental mechanisms of protein-protein association, recognition, and binding of utmost importance. The conformational selection theory claims that all protein conformations pre-exist within the dynamic apo ensemble, including the conformation of the bound state, possibly only as a high-energy state. As the Protein-Protein Binding in Two Steps substrate recognizes and binds to this conformation, the removal of the stable complex from the apo equilibrium leads to a shift of the populations toward the bindingcompetent conformation [4,5,6]. Motivated by cases in which an interplay of induced fit and conformational selection seems to guide protein binding [7], Csermely et al [8] incorporate the induced fit mechanism into an extended conformational selection model, describing shifts of the energy landscapes as part of an adjustment process caused by mutual interactions [7,8,9]
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