Many biomolecules function by interacting with other biomolecules in the cell. Some examples are:
We will analyze the interaction between tRNA and aminoacyl-tRNA synthetase. Aminoacyl-tRNA synthetases are a family of enzymes that attach the correct amino acid to each tRNA molecule. For every amino acid, there is at least one tRNA molecule and one aminoacyl-tRNA synthetase. Correct pairing of amino acids and tRNA molecules is critical for correct translation of the genetic code. Any mistake in matching an amino acid to its tRNA will result in a mutant protein.
How does each of the aminoacyl-tRNA synthetases know which amino acid should be paired with which tRNA? In the scientific jargon: how does aminoacyl-tRNA synthetase recognize the right amino acid and the right tRNA? The answer to this molecular recognition problem may lie in specific interactions that allow each aminoacyl-tRNA synthetase bind only a specific amino acid, and a specific tRNA molecule. Scientists are trying to understand the principles of molecular recognition because this allows to design novel molecules that bind and selectively act on desired biological molecules.
Reinitialize PyMOL and load the structure file 1F7U.pdb. This structure file contains two macromolecules, the protein (chain a), and tRNA (chain b). You want to separate these into two individual objects for a further study. Create object "protein" from chain A, and object "tRNA" from chain B and color these uniquely. Save each of these objects as PDB files protein.pdb and trna.pdb in your directory via Save Molecule under File menu.
How does each of the aminoacyl-tRNA synthetases know which amino acid should be paired with which tRNA. The answer may lie in specific interactions that allow each aminoacyl-tRNA synthetase bind only a specific amino acid and a specific tRNA molecule. However, coloring objects by arbitrary colors tells us little about interactions.
Think what kind of interactions are likely to occur between the polyanionic tRNA and a protein? Recall that opposites attract! Wouldn't it be nice if one can quickly check if the recognition surface of tRNA synthetase is strongly positively charged so it could well bind to tRNA?! One could, for example look for positively charged amino acids on the surface. Try to find some positively charged amino acids in the tRNA binding region of the protein.
It turns out that there is a better way to address the question of electrostatic attraction between macromolecules. Recall from your physics classes that a point charge creates an electric field around it, and the electric field is related to the charge density via Gauss's law. We could thus calculate the electric field around the protein, but the electric field is a difficult property to visualize because it is a vector quantity. A related property, the electric potential (or electrostatic potential) is easier to visualize because it is a scalar quantity, and can be mapped onto the protein surface. The electrostatic potential can calculated via Poisson's equation; such a calculation can be performed with the Adaptive Poisson-Boltzmann Solver that is installed as a Plugin in PyMOL.
Reinitialize PyMOL and load the file protein.pdb that you had saved earlier. Launch the APBS Tools plugin and hit Set Grid. The Poisson's equation is solved numerically using three-dimensional grid; the grid points are determined automatically based on the size of your protein. Not hit Run APBS. The program will first try to assign charges to all the atoms, but because of problems with this PDB file, or presence of unusual moieties, there are a number of atoms that are not properly recognized. PyMOL had created an unassigned selection of these; go ahead and use the Actions menu to remove atoms that were recognized. Now reRun APBS. The calculation takes good amount of time but if everything goes well, you should see a message "Thanks for using APBS!" in the Unix shell used to launch PyMOL.
Now go to Visualization and hit Show to display the surface. Your computer needs to be equipped with a decent graphics card to accomplish this step. This is indeed a highly charged protein, and to enhance the visual analysis alter the Low and High values to -16 and 16, respectively. Look at the resulting surface. The white areas are due to neutral, usually hydrophobic residues. The red regions indicate a negative potential due to negatively charged residues; the blue indicates a positive potential due to positively charged residues. Rotate the molecule and notice that one side is rather blue and the other side is more red/white. What do you think is the side that binds tRNA?
Load the tRNA from the file trna.pdb that you saved earlier. Did it bind where you were expecting it to bind? Show the tRNA object as a cartoon and type set cartoon_ring_mode, 1 to emphasize the purine and pyrimidine rings.
Assignments:
The structure and function of tRNA synthetase are further described in a Chime-based tutorial from Kenyon College. Please note that you need a Chime plugin to see this structure. The plugin is not installed on Linux computers.