Computer-Aided Drug Design Homework: 3. Computer Visualization
1. From Lead to Drug Candidate
This assignment is based on a recent and ongoing development of anti-cancer drugs that target a particular protein kinase. As discussed in a paper "Protein kinases — the major drug targets of the twenty-first century" , protein kinases are difficult targets because many inhibitors that mimic ATP show low specificity. Some specific protein kinase inhibitors have reached the market successfully (e.g. Gleevec), and several are in clinical development. However, other promising specific protein kinase inhibitors (e.g. Iressa), have failed to live up to their promise.
Target structure-based drug design is widely used in design of specific kinase inhibitors because many kinases are small proteins that were relatively easy to crystallize. The design of protein kinase inhibitors is further discussed in a paper "Protein Kinase Inhibitors: Insights into Drug Design from Structure".
Analyze the structure of a protein kinase with a lead compound (structure on the right, the binding constant 0.8 micromolar) and modify the lead to create two molecules that you think will bind better. You may be able to computationally test the binding of your modifications against the solutions that the company scientists came up with in the following weeks. You are allowed to use any resource and approach you deem suitable. Perform the following tasks:
- Download the structure of the protein and the docked lead compound. Open both in a suitable visualization program and examine the binding pocket and the position of the lead compound.
- Based on your own ideas, and readings of protein-kinase inhibitor design literature, outline your strategy to optimize the lead compound.
- Closely examine how the lead compound is bound to the target. Take advantage of 3D stereo capabilities offered by the Linux workstations. Identify limitations of the current lead compound. Create an image of the bound lead that emphasizes at least one of these limitations.
- Starting with the structure of the lead, create two new molecules that you expect to bind better. Notice that the active site for the protein kinase is rather large as it must accommodate both substrates; you are not expected to fill the whole active site with your drug! You can use program of your choice. For example, you could edit the lead in MOLDEN and write it out as PDB file. Simple editing, such as addition or deletion of methyl groups can be also readily done with PyMOL. Students who are familiar with SYBYL's editing capability might find this program a good choice for creating modified structures. Explain in detail the reationale behind making such modifications.
- Superimpose the modified molecule with the original lead in PyMOL or Chimera. To manually superimpose the modified drug candidate with the bound lead in PyMOL, you have to protect the protein and lead from movement, and deprotect the modified compound. 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". Students who are familiar with SYBYL's might find this program a good choice for superimposing structures.
- Create two images showing each of your modified structures superimosed to the lead in the active site. Show the protein cavity as a surface colored by the electrostatic potential. Use transparent surfaces if you want to emphasize specific interactions with protein that your modification allows for.
- Provide the chemical structure of your two modifications as usual chemical drawings (e.g. IsisDraw).
2. Fragment Based Design
The structure of acetylcholinesterase (2C58) that you analyzed in the previous tutorial contained more than one bound ligand. Some of the bound these ligands were bound in the active site region, others were further away. In this task, you will use the structures of the two small ligands that were bound in the active site region and design a larger ligand that will hopefully display tighter binding. Perform the following tasks:
- Discuss how is it possible that a protein that is target of some of the most lethal chemical warfare agents is being seriously considered as a target for anti-Alzheimer's drugs? What structural features should a drug designer avoid in order to minimize toxicity of acetylcholineesterase binding molecules?
- Based on the available literature, write a brief description about the catalytic mechanism of acetylcholinesterase. Create one illustration either with chemical drawing software (such as IsisDraw, or jChemPaint) or with molecular visualization software that depicts the catalytic mechanism of this enzyme.
- Examine the information available in the PDB website to identify which ligands bind to this enzyme and where do they bind. Locate these ligands in the structure and create a picture showing both of the bound ligands in the binding pocket.
- Following the fragment-based strategy, design one larger molecule that appropriately occupies the positions of the two small ligands. Manually place this molecule in the active site. It is likely that you have to manipulate the geometry of the larger ligand to some degree to avoid steric clashes. This can be achieved in PyMOL via "sculpting", followed by partial quantum mechanics minimization in MOLDEN (set all dihedrals to constant in Z-matrix". Optionally, students who are more familiar with biomolecular modeling may consider minimizing the structure of the bound ligand within the active site with a program they are familiar with.
- Create an image showing how the larger drug binds to the active site. Provide the chemical structure of the molecule you design.