Site-specific binding affinity of Eu(III) towards Ca-binding protein calmodulin: A combined spectroscopic and theoretical study


Site-specific binding affinity of Eu(III) towards Ca-binding protein calmodulin: A combined spectroscopic and theoretical study

Tsushima, S.; Samsonov, S.; Drobot, B.; Raff, J.; Komeiji, Y.; Mochizuki, Y.

Molecular modelling of actinide interaction with large biomolecules by quantum chemical calculations is restricted by request for huge computational resources. We challenged this problem by applying Fragment Molecular Orbital (FMO) method. In FMO, the molecular system is partitioned into small fragments and each fragment and fragment pair is subjected to self-consistent field calculations which drastically reduces computational cost.1 We are upgrading FMO program Abinit-MP 2 to implement f orbitals. Here, interaction of Eu3+ with Ca-binding protein calmodulin (CaM) was studied. Calculations were performed in the following way. Using the crystal structure of Ca2+-bound CaM, all four Ca2+ ions were replaced by Eu3+, protonation state of Eu-CaM was adjusted, and 12 Na+ were added for neutralization. The structure was immersed in a TIP3P water bath of 8 Å thickness around Eu-CaM and submitted to 100 ns molecular dynamics (MD) run. Structure at each 1 ns was collected (100 samples), waters were stripped off to 4 Å coverage, and the structures were used in FMO calculations at MP2 level. The statistical average of inter-fragment interaction energy (IFIE) was calculated. Average coordination number of Eu in Eu-CaM during 100 ns MD run is between 8.9 and 9.1. Eu sits in the same binding site as Ca but with increased bidentate coordination. Additionally, increased water coordination is observed. In EF Hand 1, 2, and 4, there is an average of 1.7 to 1.9 coordinating waters to Eu whereas in Hand 3 there is an average of 2.9 waters. This result perfectly matches with previous spectroscopic findings where 2 waters was found at sites 1 and 2 and at either site 3 or site 4, with 3 waters at the remaining site.3 When we compare four metal-binding sites in Eu-CaM, the IFIE between Eu3+ and the corresponding binding site are overall similar among three of the four binding sites, namely EF Hand 1, 2, and 4. On the other hand the IFIE for Hand 3 is clearly smaller pointing to that Hand 3 is the weakest binding site. The reason for this is clear; EF Hand 3 carries only three negatively charged residues, whereas the other motifs have four of them. Consequently, Eu binding in EF Hand 3 exhibits relatively larger fluctuations compared to other binding sites which causes structural disorder to Eu-CaM. We also performed titration and Eu luminescence lifetime measurements which are found to be consistent with MD and FMO results.

  • Lecture (Conference)
    Actinides 2017, 09.-14.07.2017, Sendai, Japan

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Publ.-Id: 25284