Step 1

Formulating Your Hypothesis As A ProcessDB Diagram

Biological diagrams represent molecules and molecular complexes in a variety of physiological places or locations. These diagrams also indicate the processes that transport molecules from one location to another, the biochemical processes that transform molecules, and the binding processes that form homo- or heteromultimers from individual molecules. All of these features are easily captured in ProcessDB diagrams.

Add the physiological places to your diagram. Click the Places tab in the database pane. Click the new place icon and enter the name of the place you want to add to your model. Places are the physiological locations of your molecules and molecular complexes. Examples are cytoplasm, plasma membrane, ER lumen, extracellular space, blood plasma. A place is the answer to the question, “Where is this molecule located in the system I am studying”? You can add all your places immediately or add them as you need them. Notice that molecules and complexes may be present in more than one place.

If the place name you chose is already in the ProcessDB database you can use it to build your own diagram; if it is not already in the database, your name will be added to the place list. In either case your place will be highlighted in the place list and you can now drag it onto the diagram pane. For example, if you type Cytoplasm and then drag the name from the database pane to the diagram pane you will see a yellow rectangle named Cytoplasm added to your diagram.

Hint: It is sometimes useful to define places that you might not ordinarily think of as places. For example, if you want to model a molecular complex whose composition changes with time, it can be very helpful to define that complex as a "place". This allows you to add the various molecular constituents to that complex and have the relative abundances of these constituents change with time. A good example of this is a plasma lipoprotein, like the famous HDL or LDL. These are the vehicles carrying lipids in the blood. They are composed of a phospholipid-cholesterol surface monolayer and a neutral lipid (triglyceride and cholesteryl ester) core. By treating LDL as a ProcessDB place it's easy to account for changes in lipoprotein composition with time.

Suppose your theory or hypothesis includes a biochemical reaction. As a specific example we take the degradation of cAMP to AMP. Click the Molecules and Complexes tab. Type cAMP in the search box on the Molecules and Complexes tab. Notice it’s already in the database and ready for you to use. Drag cAMP into the light yellow Cytoplasm place. By the way, if cAMP had not been in the database you could have used the new molecule icon () to add it. Now your screen looks like this:

Notice that the tree structure pane in the lower left of the screen now shows a state called cAMP in Cytoplasm. A State in ProcessDB is a molecule or molecular complex in a particular place. Next find AMP in the molecule list either by scrolling or by typing AMP in the search box. Then drag AMP into the cytoplasm. This yields:

Now we are ready to add the process that converts cAMP to AMP. In ProcessDB all processes and control mechanisms are added to the diagram using the Connect tool () on the diagram pane toolbar. Select the connect tool. Now drag from the substrate to the product. A dialog appears with a default name for this process. You can accept this name or type cAMP phosphodiesterase to rename it. Next click one of the layout buttons () to produce a hierarchical, symmetric or orthogonal layout of the current diagram. ProcessDB takes care of layout automatically even for extremely complex diagrams. Here’s the result:

Suppose you also wish to represent a transport process such as cAMP secretion into the extracellular space. On the place tab scroll or search for Extracellular space. Drag this place onto the diagram. Then drag, from the Molecules and Complexes tab, cAMP into the extracellular space. Finally select the Connect tool and drag from cAMP in Cytoplasm to cAMP in Extracellular space. Click one of the Layout tool buttons to arrange the result neatly.

Suppose you want to add binding of cAMP to a cyclic AMP receptor in the plasma membrane. In the Places tab of the database pane scroll or search for plasma membrane and drag this place to the gray region between the extracellular space and the cytoplasm. This placement is not in any way necessary but it does make the diagram more like the cell.

In the Molecules and Complexes tab scroll or search for cAMP receptor. Drag this molecule to the plasma membrane. Save your work to the database by clicking the Save icon () and click a Layout button.

Now we need to create the molecular complex consisting of cAMP and the cAMP receptor. To do this click the New complex button () on the Molecules and Complexes toolbar. This opens a blank list to which you drag the molecules you want in the new complex. Type cAMP in the search box so that both cAMP and cAMP receptor are made available in the database pane. Then drag each molecule to a line in the list of molecules in the new complex.

Tip: If your complex involves, say, insulin and insulin receptor, we recommend the automatic name insulin:insulin receptor. This default name will be assigned automatically; there is no need to enter the default name in the Complex Name field. You may want to modify the default name if your complex involves so many molecules that its default name becomes unwieldy. In this case you can replace the long default name with an abbreviation by selecting the entire default name in the Complex name field and typing over it with your abbreviation.

Click Save at the bottom of the new complex molecule list. If the stoichiometry was different from 1:1 you could change the numbers in the second column to reflect this. The new complex is added to the Molecules and Complexes list and is immediately highlighted. Drag it to the plasma membrane.

Last, select the Connect tool again. Since the binding reaction involves two molecules, adding it to the diagram requires two steps. First drag an arrow from cAMP in Extracellular space to cAMP:cAMP receptor in plasma membrane.

Second, drag an arrow from cAMP receptor in plasma membrane to the ARROW labeled cAMP receptor binding. This opens a dialog in which you choose the role to be played by cAMP receptor in this process.

Select Start or Substrate to indicate that cAMP receptor in plasma membrane is an additional reactant in this process. Click a layout button and then the Save icon to commit your work to your database.

To make the binding process reversible, use the Connect tool and drag an arrow from cAMP:cAMP receptor to cAMP receptor. Call this cAMP receptor unbinding. Then drag an arrow from the cAMP receptor unbinding arrow to cAMP in Extracellular space. This suffices to indicate that cAMP is a second product of the unbinding reaction. Click orthogonal layout, and save your work.

This diagram now contains examples of all three classes of basic biological processes. cAMP phosphodiesterase is an example of a chemical reaction. cAMP secretion is an example of transport, and cAMP receptor binding is an example of binding.

ProcessDB Diagrams can also indicate other roles for the states in your diagram. These include all the roles listed in the connect tool dialog box shown earlier and re-shown here:

Exercise: Add the enzyme cAMP phosphodiesterase to the diagram. Drag an arrow from it to the cAMP phosphodiesterase process. When the Role dialog appears, select Enzyme and OK. Click incremental layout (it's the third icon from the right on the Diagram tool bar) and then save your work by clicking the Save icon (it's on the Main toolbar just below the View menu).

This model is now ready for Step 2 in which the qualitative pathways are augmented with quantitative rate laws.

Tip: If you are already impatient with the pace of this tutorial, and you just want to see what sort of output ProcessDB will produce, you might want to explore Run Initial Simulation on your model's context menu. Almost everything on the ProcessDB screen has a context menu that is accessed by right clicking.