1. Development of Software Package for Determining Protein Titration Properties By Kaila Bennett, Amitoj Chopra, Jesse Johnson, Enrico Sagullo

2. Overview This talk will address how our group proposes to develop a stand-alone software package capable of elucidating protein titration characteristics using the thermodynamic cycle Adapted from Morikis et al

3. Background It is well known that proteins are highly dependant on the pH of the environment Functional groups within the protein can extract or release protons This proton transfer phenomena lead to the changes in ionizable charge groups: Aspartic acid Glutamic acid Arginine Lysine Histadine Charge amino acids have titratable side chain groups These titratable groups have characteritistic pK a ’s

4. Background (pK a ) The thermodynamic cycle can be used to calculate pK a Within each pK a resides an intrinsic and apparent pK a The intrinsic pK a describes the ionization process of a specific ionizable group when all other ionizable groups are held constant (Morikis et al ). The apparent pK a relates the intrinsic pK a and the free energies between charged residues Cycle adapted by Morikis et al

5. Background To extract titration properties we will be writing scripts using the programming language, R R programming language used for statistical computing and graphics Using R, we will be able to integrate free energy calculations using Adaptive Poisson-Boltzmann Solver (APBS)

6. Purpose of the Project Using R programming, we want to write a function that converts PDB files to PQR files. Once the program is complete, we will use APBS software to calculate free energies and electric static potentials. These calculations will then aid us in writing a program that will calculate intrinsic and apparent pK a values

7. Objectives Winter Get acquainted with R programming language Workshops with Dr. Girke Tutorial help from Chris Kieslich Incorporation of APBS Allows us to calculate the intrinsic pK a values Calculates Free energies Have software able to take any protein database files (pdb) and calculate intrinsic pK a to generate titration curves Examples of titration curves generated in R

8. Objectives Spring Statistical approximation for the calculation of apparent pK a values using the intrinsic values Clustering Monte Carlo Add scripts to print out titration curves for each and all ionizable amino acids Calculate protein stabilities and binding free energies Example of clustering in R

9. Methodology Winter

10. Methodology Spring

11. Budget As of now, we do not foresee any expenditures due to the fact that we are using all free software

12. Relevant Equations

13. Conclusion The goal of this project is to create a local downloadable software package that will be able to calculate the titration properties for any protein The software will first calculate the intrinsic pK a values of each residue which will then be correlated into the apparent pK a values The software will work to generate titration curves for easy to view analysis The software will integrate already defined software such as APBS to in essence create a user-friendly convenience package. It will be a stand-alone software free and available for anyone to use

14. Acknowledgements Dr. Dimitrios Morikis Chris Kieslich Dr. Thomas Girke Dr. Jerome Schultz Hong Xu

15. References 1. Trylska, Joanna. "View Continuum Molecular Electrostatics, Salt Effects, and Continuum Molecular Electrostatics, Salt Effects, and Counterion Binding—A Review of the Poisson– Boltzmann Counterion Binding—A Review." Wiley InterScience 28.2 (2007). Print 2. Antosiewicz, Jan M. "Protonation Free Energy Levels in Complex Molecular Systems." Wiley InterScience 89.4 (2007). Print 3. Gilson, Micheal K. "INTRODUCTION TO CONTINUUM ELECTROSTATICS, WITH MOLECULAR APPLICATIONS." Editorial. 13 Jan. 2006. Print 4. Morikis, Dimitrios. "Molecular thermodynamics for charged biomacromolecules." Fluid Phase Equilibria (2006). Print 5. Nielsen, Jens. "Analyzing Enzymatic pH Activity Profiles and Protein Titration Curves Using Structure-Based pK a Calculations and Titration Curve Fitting." Methods in Enzymology. Print.

16. Questions