Interactions with the Binding Site
Specific, high-affinity binding typically requires that the bound drug makes several favorable polar contacts with the receptor. Binding affinity, or drug selectivity, can be enhanced by optimizing these interactions by slightly modifying the lead compound. The binding affinity is traditionally calculated using highly simplified force fields in a process called docking. However, when only a few lead modifications are considered, one could use more accurate, but also more time consuming quantum mechanical calculations to assess drug-receptor interaction enthalpies.
Tutorial: Interaction Energy of Chorismate Mutase with its Transition State
Your task is to calculate the interaction energy between the active site of chorismate mutase, and the transition state of chorismate-to-prephenate reaction. To perform these calculations:
- Start with the active site that has been extracted out from the X-ray structure 1ECM (file 1ecm_asite.mol. This structure contains a dianionic oxa-bicyclic inhibitor. Open this structure with PyMOL.
- Create a structure of dianionic transition state. For now, use a PM3-optimized structure from file chr_da_tsen_pm3.mol. When you work with other molecules as a part of the assignment, you need to create such optimized dianionic structures from available monoanionic structures yourself. PM3 optimization is sufficient for our purposes.
- Open the structure file for the dianionic transition state in the same PyMOL window that has the active site. Zoom out until you see both the active site and the transition state. Change the clipping plane such that both structures are well visible.
- Superimpose the transition state with the inhibitor in PyMOL. To manually superimpose the modified transition state, you have to protect the active site from movement, and deprotect the transition state. The protect and deprotect commands are also available from the right-side tool-box under Actions: movement. You can translate and rotate the deprotected objects by holding down the SHIFT key while in the "Mouse Editing Mode". You can translate and rotate both structures when you release the SHIFT key. Students who are familiar with SYBYL's might find this program also a good choice for superimposing structures.
- Once a close superposition is achieved, refine the superposition using the Pair Fitting Wizard in PyMOL. When using this Wizard, click first on an atom (mobile atom) the transition state, then on a matching atom (target atom) in the inhibitor. Pick six pairs based on the atoms in the six-membered ring with only carbon atoms and Fit based on these pairs.
- Hide the transition state and delete all inhibitor atoms from the active site. Deletion of atoms can be easily achieved by first selecting inhibitor Atoms in the "Mouse Viewing Mode", and then executing
Remove atoms action from the Actions: side-menu. Make sure that you are deleting atoms only from the (sele) (or pk1 object), not from the 1ecm_asite or chr_da_tsen_pm3 object.
- Combine the superimposed transition state, and the empty active site into one object by typing create 1ecm_asite_ts, 1ecm_asite or chr_da_tsen_pm3 into PyMOL command line.
- Save the combined active-site-transition state object 1ecm_asite_ts via the Save Molecule menu as a mol file.
- Save the empty active site object 1ecm_asite via the Save Molecule menu as another mol file.
- Determine the net charge of the empty active site, and the complex by visual inspection
- Open the isolated TS structure in MOLDEN and perform a Single Point Energy calculation using the PM3 model.
- Open the empty active site structure in MOLDEN and perform a Single Point Energy calculation using the PM3 model.
- Open the active site-TS complex structure in MOLDEN and perform a Single Point Energy calculation using the PM3 model.
- Calculate the interaction energy between the transition state and the active site
Assignments
- Find the lowest-energy geometry for the CO2 dimer using AM1, PM3, PM6, HF/6-31+G(d), and MP2/aug-cc-pVTZ methods. This task involves building the CO2 as a monomer, and minimizing the monomer followed by building an initial guess structure (T-shaped or slipped-parallel?) for the CO2 dimer and minimizing it. You may use MOPAC for semiempirical calculations. Gaussian is appropriate for HF and MP2 minimizations. When using Gaussian, you need to manually specify the aug-cc-pVTZ basis set instead of STO-3G, and remove the 6D 10F options from the second Keyword line. Gaussian and many other quantum chemistry programs give energies in Hartree units; you can convert from Hartrees to kcal/mol by multiplying with 627.51. (3 pts)
- Read the paper "Thermodynamic and Structural Effects of Conformational Constraints in Protein-Ligand Interactions. Entropic Paradoxy Associated with Ligand Preorganization". Your goal is to test the hypothesis that the unusual findings discussed by the authors arise from differential solvation of flexible and constrained analogues. Each student in the course will calculate the aqueous solvation free energy for a pair of molecules (e.g. the first student will do molecules 5 and 11 in the paper). You are encouraged to work together when setting up these calculations. Because these are rather large molecules, use reasonable quantum chemistry methods with modest basis sets. For example, HF/3-21G optimization or B3LYP/6-31+G(d,p) single point might be all right choices but the optimization at the MP2/aug-cc-pVTZ level would certainly take too long. Each student can pick his/her favorite quantum mechanical method but the two molecules in a pair should be always treated the same way. For the sake of efficiency, omit the phosphate moiety, i.e. just use the phenyl group at position 5. Collect your results and discuss if the unusual finding in this paper can be rationalized by differential solvation of compounds 5 and 11. Submit one answer with your results and discussion as a group; everybody will get the same number of points for this task. (3 pts)
- Following a protocol similar to that in the tutorial, calculate the interaction energy between the active site of chorismate mutase, and the oxa-bicyclic inhibitor, the methylene-bridged bicyclic analog, the molecule that you proposed as an inhibitor of chorismate mutase, and the methyl analog of oxa-bicyclic inhibitor that I proposed as part of the first homework (see the key). Briefly discuss main limitations of calculating the interaction energy with the approach outlined in the tutorial. How would you change the protocol in order to get more meaningful results? (4 pts)