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Enzyme Active Sites
This module includes models of three serine proteases: chymotrypsin, trypsin, and elastase. The goal of
the module is to give students a grasp on the size and shape of enzymes and the structural features that
are important in creating an active site. We envision use of the models in a lab setting, with groups of 4-5
students working with each model, and trading models throughout the session. We expect that they will
be most effective if the students are allowed to explore them directly.
Guide for Exploration
1. Get familiar with the model:
a. Start with a spacefilling model. Each atom is represented by a sphere--larger spheres for carbon,
nitrogen, oxygen and sulfur and smaller spheres for hydrogen. Make an estimate of the number
of atoms in the enzyme.
b. The coloring scheme highlights the chemical characteristics of the atoms. Carbon atoms are
colored white. Oxygen is colored red--bright red when charged and pink when neutral. Nitrogen is
colored blue--bright blue when charged and light blue when neutral. Sulfur is colored yellow.
Hydrogen atoms are colored the same color as the atoms they are attached to. Find examples of
each atom type.
c. Notice the packing of sidechains to give a virtually seamless surface
d. Notice the presence of many small pockets and grooves. Are these filled with water molecules in
the crystal structure?
2. Find amino acids:
a. Find arginine, lysine, glutamic acid and aspartic acid (the two acidic amino acids are hard to
distinguish from one another!) Notice the distribution of charged amino acids on the surface. Are
there salt bridges?
b. Find asparagine/glutamine, serine/threonine and histidine. Notice the distribution of polar amino
acids on the surface. Can you find examples of hydrogen bonds between groups?
c. Find phenylalanine, tyrosine, tryptophan and methionine. Notice the relative lack of these large
hydrophobic amino acids on the surface. Are these involved in stabilizing interactions?
d. Look for disulfide bridges between cysteines.
3. Find the active site:
a. Look for a groove that binds to protein chains
b. Look for the specificity pocket, which is different in each: a large pocket that binds to aromatic
sidechains in chymotrypsin, a deep pocket with an acidic group at the bottom in trypsin, and a
small pocket in elastase.
c. Look for an activated serine that performs the cleavage, and the charge relay system of serinehistidine-aspartate.
d. Look for two backbone amide nitrogen atoms pointing into the active site that stabilize the
negative charge on the protein chain that is being cleaved. This has been termed the "oxyanion
hole".
e. Take a minute to explore the specificity pocket, and how it is constructed. Notice how the amino
acids around the pocket form the specific shape. Notice that it is the only pocket of this size in the
protein: somehow the protein is holding this space open with the right shape.
4. Compare the backbone model of the protein with the spacefill model of the same protein.
a. Backbone models highlight secondary structure: alpha helices are colored yellow and beta sheets
are colored red. The N-terminus is blue and the C-terminus is red.
b. See if you can orient the backbone and spacefill models.
c. What features are best identified in the spacefill model? In the backbone model?
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