(749b) Forcefield_Ptm and Forcefield_Ncaa: Platforms for Simulating Modified Amino Acids in AMBER That Can Reproduce Experimental Thermodynamic Properties

We describe the development and testing of two new ab initio derived atomistic forcefields, Forcefield_PTM¹ for post-translational modifications and Forcefield_NCAA² for unnatural/non-canonical amino acids for the AMBER (http://ambermd.org) molecular dynamics suite of tools. The parameters were made to be compatible with the AMBER forcefield ff03³. Forcefield_PTM addresses the most frequently occurring post-translational modifications in nature⁴, and Forcefield_NCAA addresses 147 commercially available and potentially druggable non-canonical amino acids including β- and N-methylated amino acids. The developed parameter libraries include charge parameters calculated using the RESP fitting approach⁵ and torsion parameters optimized to enable the modeling, simulation, and design of modified protein structures in combination with AMBER ff03³. The charges were optimized using the B3LYP/cc-pVTZ//HF/6-31G** level of theory. The forcefields have been optimized to reproduce the quantum mechanically calculated electrostatic and torsion potential energies along key barriers.

Although the forcefields have been trained to reproduce quantum chemically calculated electrostatic potentials and energies, they have been tested on multiple orthogonal experimental metrics including binding free energies of BCL9/Beta-catenin inhibitors⁶, hydration free energies of side-chain analogs⁷, and to separate active/inactive inhibitors of Complement activation based on IC50 values to high accuracy. The forcefield can rank binding free energies with an R² of 0.76, separate active/inactive inhibitors of Compstatin to 80% accuracy, and rank hydration free energies of non-canonical amino acid side-chain analogs with an RMS of 2.53 and MAE of 2.10 kcal/mol and an R² of 0.68. They have additionally been used in conjunction with replica-exchange to fold a post-translationally modified protein from scratch. The forcefields are now actively being used to design new inhibitors of HIV⁸ and improper Complement activation⁹ with some recent success.

The parameters are freely available to academic researchers at http://selene.princeton.edu/FFPTM and http://selene.princeton.edu/FFNCAA to aid in studying their own systems complete with an interactive web interface to introduce modified amino acids into proteins. We describe the development, challenges, validations, and applications of our forcefields and the plans for their future development. The forcefields allow one to incorporate customized amino acids site-specifically into design applications with control over size, length, shape, and electrostatic properties.

 

References:

1.         Khoury GA, Thompson JP, Smadbeck J, Kieslich CA, Floudas CA. Forcefield_PTM: Ab Initio Charge and AMBER Forcefield Parameters for Frequently Occurring Post-Translational Modifications. J Chem Theory Comput 2013;9(12):5653-5674.

2.         Khoury GA, Smadbeck J, Tamamis P, Vandris AC, Kieslich CA, Floudas CA. Forcefield_NCAA: Ab Initio Charge Parameters to Aid in the Discovery and Design of Proteins and Peptides with Unnatural Amino Acids and Their Application to Complement Inhibitors of the Compstatin Family. ACS Synthetic Biology 2013(In Press):DOI:10.1021/sb400168u.

3.         Duan Y, Wu C, Chowdhury S, Lee MC, Xiong G, Zhang W, Yang R, Cieplak P, Luo R, Lee T, Caldwell J, Wang J, Kollman P. A point-charge force field for molecular mechanics simulations of proteins based on condensed-phase quantum mechanical calculations. J Comput Chem 2003;24(16):1999-2012.

4.         Khoury GA, Baliban RC, Floudas CA. Proteome-wide post-translational modification statistics: frequency analysis and curation of the swiss-prot database. Scientific Reports 2011;1(90).

5.         Bayly CI, Cieplak P, Cornell W, Kollman PA. A well-behaved electrostatic potential based method using charge restraints for deriving atomic charges: the RESP model. The Journal of Physical Chemistry 1993;97(40):10269-10280.

6.         Kawamoto SA, Thompson AD, Coleska A, Nikolovska-Coleska Z, Yi H, Wang S. Analysis of the interaction of BCL9 with beta-catenin and development of fluorescence polarization and surface plasmon resonance binding assays for this interaction. Biochemistry (Mosc) 2009;48(40):9534-9541.

7.         Khoury GA, Bhatia N, Floudas CA. Hydration free energies calculated using the AMBER ff03 charge model for natural and unnatural amino acids and multiple water models. Submitted 2014.

8.         Khoury GA, Tamamis P, Smadbeck J, Szekely Z, Sinko PJ, Floudas CA. Rational Computational and Experimental Design of Peptides Targeting HIV-gp41 with Non-canonical and Cross-linked Amino Acids and Insights into their Mechanism of Inhibition In Preparation 2014.

9.         Floudas CA, Khoury GA, Smadbeck J; The Trustees of Princeton University, assignee. Computational Framework and Parameters for the Discovery of Therapeutic Proteins and Peptides with Post-Translational Modifications and Non-Canonical Amino Acids. United States of America. 2013 May 17, 2013.