Ation, or a minimum of a pointer towards how this should be completed. Authors’ response: We are delighted to determine that Reviewer appreciated the scale with the trouble that the object of this study has set for theoretical calculations. We thank the reviewer for his incredibly valuable comments. We agreed and have taken into account all of them with the single exception in the one particular that had been marked as an error by the Reviewer. We nevertheless think that we have used a suitable criterion for the salt bridges in our evaluation. Figure 1a and b, the necessity of which has been questioned by the Reviewer in the comment (34), show how our final model fits inside the EM density. In the revised manuscript we give some hints on how the functional consequences from our model could beShalaeva et al. Biology Direct (2015) 10:Web page 26 ofvalidated by mutating the acidic residues of Apaf-1. Naturally, we hope to determine a well-resolved crystal and or cryo-EM structure on the cytochrome cApaf-1 complex in the near future.More filesAdditional file 1: Figures S1 and S2. Figure S1. Backbone coordinates RMSD heat maps for WD domains of Apaf-1 in complex with cytochrome c during MD simulation. Figure S2. Conservation of negatively charged residues within the WD domains of Apaf-1 homologs. Further file two: The PatchDock’ model structure right after power L-Cysteic acid (monohydrate) Autophagy minimization. This is the structure obtained after manual editing of PatchDock-predicted model and power minimization. The PatchDock’ model shows probably the most quantity of salt bridges involving functionally relevant cytochrome c residues and remained steady throughout molecular dynamics simulations. Further file three: Original EM-fitted model structure [PDB:3J2T] [25] just after energy minimization. Further file 4: The ClusPro-predicted model structure after power minimization. Additional file five: The PatchDock-predicted model structure soon after energy minimization. Further file 6: The first ZDOCK-predicted model structure immediately after energy minimization. Further file 7: The second ZDOCK-predicted model structure right after power minimization. Abbreviations Apaf-1: Apoptotic protease activating element 1; CARD: Caspase activation and recruitment domain; Cryo-EM: Cryo-electron microscopy; Etc.: Electron-transfer chain; MD: Molecular dynamics; NBD: Nucleotide-binding domain; ROS: Reactive oxygen species. Competing interests The authors declare that they’ve no competing interests. Authors’ contributions DNS performed molecular modeling and MD simulations, analyzed the data, at the same time as wrote the initial draft of the manuscript, DVD performed the sequence analysis of cytochrome c, MYG performed the sequence 7α-Hydroxy-4-cholesten-3-one In stock evaluation of Apaf-1 and contributed to the writing the manuscript, AYM made the study, interpreted the data, and wrote the final version in the manuscript. All authors read, edited and authorized the final manuscript. Acknowledgements The authors are grateful to Prof. V.P. Skulachev for drawing their consideration towards the prospective key role of the residues of Apaf-1 within the formation of an apoptosome. The investigation of your authors was supported in aspect by the Osnabrueck University, Germany in addition to a fellowship from the German Academic Exchange Service (DNS), grants from the Russian Science Foundation (1440592, AYM, molecular modeling of apoptosome formation, and 1400029, DVD, AYM, phylogenomic evaluation of cytochrome c), by the Development Plan in the Lomonosov Moscow State University, Russia (access for the supercomputer facility), and by the Intramural Investigation System of t.
