Enzyme Mechanisms - Serine Proteases
Part VII
Part VII. Conclusion for the Mechanism of Catalysis by Serine Proteases.
Three key AA side chains of serine proteases (Trypsin, Chymotrypsin, and Elastase) which are Asp-102, His-57 and Ser-195 worked together in a concentrated way to catalyze hydrolysis of amides (proteins/peptides) and ester. Finally, you need to challenge the hypothesis for the enzyme catalyzed reaction mechanism. Today that is easily done by designing a mutant enzyme.
Designing a mutant enzyme with one AA change:
1. Clone a gene for the enzyme
2. Get a bacterium to make the cloned enzyme in active form
3. Locate a key AA and change the gene so it puts a different AA in its place.
Figure 18. Schematic diagram of the construction of a mutant enzyme.
Now have the bacteria make the mutant protein for Chymotrypsin with Ala at position 195. Determine if can catalyze the reaction- if not then Ser-195 is indeed a key AA.
One practical application of learning details of enzyme catalytic mechanisms is the design of enzymes to catalyze reactions for which no enzyme exists.
©Wilbur H. Campbell, 1995; wcampbel@mtu.edu