Abstract
Computational investigations into the structure and function of metalloenzymes with transition metal cofactors require proper preparation of the model, which requires obtaining reliable force field parameters for the cofactor. Here, we present a test case where several methods were used to derive amber force field parameters for a bonded model of the Fe(II) cofactor of ectoine synthase. Moreover, the spin of the ground state of the cofactor was probed by DFT and post-HF methods, which consistently indicated the quintet state is lowest in energy and well separated from triplet and singlet. The performance of the obtained force field parameter sets, derived for the quintet spin state, was scrutinized and compared taking into account metrics focused on geometric features of the models as well as their energetics. The main conclusion of this study is that Hessian-based methods yield parameters which represent the geometry around the metal ion, but poorly reproduce energy variance with geometrical changes. On the other hand, the energy-based method yields parameters accurately reproducing energy-structure relationships, but with bad performance in geometry optimization. Preliminary tests show that admixing geometrical criteria to energy-based methods may allow to derive parameters with acceptable performance for both energy and geometry.
Highlights
Transition metal ions fall in between group 1 and group 2 metal cations, which form mainly ionic bonds, and the p-block elements of the periodic table, which form bonds of more covalent character
Experimental data strongly indicate that the Fe(II) ion plays a central role in catalytic activity of EctC [1, 2], we aim to obtain as precise description of the metal site as possible by testing three parameterization methods for bonded approach: the
To check the importance of these iron-binding residues for EctC enzymatic activity, they were individually replaced by alanine via site-directed mutagenesis, and the results showed a substantial reduction in enzymatic activity: A drop of 94,6%, 87.3% and 84.9% with respect to the activity of the WT form was observed for the E57A, H92A and Y84A mutants, respectively
Summary
Transition metal ions fall in between group 1 and group 2 metal cations, which form mainly ionic bonds, and the p-block elements of the periodic table, which form bonds of more covalent character. In 2011, Hu and Ryde analyzed five different approaches (both bonded and non-bonded) for MM parameters derivation while applying them to zinc metalloproteins for testing purposes [7] They concluded, that before attempting the parameterization procedure, it is crucial to determine the future use of the parameters, as their performance will heavily depend on the nature of the metal-binding site, for example, they recommend using Norrby-Lijefors automated method for the catalytically important ions [7, 8]. In 2017, Li and Merz published a comprehensive review of the metal site parameterization methods [9] Their goal was to cover the current state of the art in one paper in order to facilitate the decision-making over which approach to choose, without testing the performance of the methods in proteins. All of the methods tested here are discussed in the 2017 review [9], but here we apply them to perform parameterization and we validate the performance of the resulting parameter sets
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