Final Report Summary - MOLDYKITA (Solvent dynamics in enzymatic catalysis: a molecular dynamics simulations and kinetic terahertz absorption spectroscopy study) The main aim of the project MOLDYKITA is to characterize by means of molecular dynamics (MD) simulations and time resolved kinetic THz absorption spectroscopy (KITA) techniques the solvation dynamics for a physiologic scenario of enzymatic catalysis, namely the interaction between a zinc metalloprotease enzyme (MMP14) and collagen-like molecules (a triple helix peptide, THP, and a single chain peptide, SCP). A particular challenge is to identify and quantify the influence of these different substrates on the water network and water dynamics and therefore to point out the functional role of substrate flexibility.The fellow, working in an interdisciplinary research team, has used kinetic terahertz absorption (KITA) spectroscopy to study protein-water dynamics during proteolysis of the two investigated systems. Notably, changes in collective enzyme–substrate–water-coupled motions have been detected, with a persistence well beyond steady state for both substrates and with a substrate-specific behavior [1]. By means of MD simulations, the fellow has provided a microscopic picture of the protein and water dynamics in the two investigated systems. In particular, she has characterized the influence of the two different substrates on the water dynamics in the catalytic domain of the enzyme in the enzyme-substrate complexes, revealing a solvent dynamical heterogeneity at the functional site: the fellow has provided evidence for a strong, substrate-specific water−protein coupling as the cause of the observed heterogeneity [2]. In addition, the fellow showed that a hydration funnel, i.e. a gradient in retardation of hydrogen bond dynamics toward the active site, characterizes both the investigated systems and that this funnel is substrate-dependent, exhibiting a steeper gradient for the more complex enzyme–collagen system (i.e. MMP14-THP) [1].Altogether, these results suggest that the long-lasting changes in protein–water dynamics reflect a collection of local energetic equilibrium states specifically formed during substrate conversion. Thus, the observed long-lasting water dynamics contribute to the net enzyme reactivity, impacting substrate binding, positional catalysis, and product release [1]. Importantly, the substrate-specific water-protein coupling suggests that the substrate flexibility plays a functional role.The novel and significant knowledge that has been gained within the MOLDYKITA project is expected to have a huge impact in the understanding of molecular recognition processes, which is of paramount importance for Physics, Chemistry and Biology. References:[1]J. Dielmann-Gessner, M. Grossman, V. Conti Nibali, B. Born, I. Solomonov, G. B. Fields, M. Havenith and I. Sagi, Enzymatic turnover of macromolecules generates long lasting protein-water coupled motions beyond reaction steady-state, Proc. Nat. Acad. Sci., USA vol. 111 no. 50 17857-17862 (2014). [2]V. Conti Nibali and M. Havenith, New insights into the role of water in biological function: Terahertz absorption spectroscopy and molecular dynamics simulations studies of the solvation dynamics of biomolecules, J. Am. Chem. Soc. 136, 12800-12807 (2014).
