TABLE OF CONTENTS Cells function by the action of enzymes. Life is a dynamic process that involves constant changes in chemical composition. For these, chemical enzymes are required. These changes are regulated by catalytic reactions, which are regulated by enzymes. That s why enzymes are called biological catalysts.
In exercise 3, we will extract the enzyme tyrosinase and study its kinetic parameters. It is only one of thousands of enzymes working in concert within cells, but it is one that readily demonstrates the main features of enzyme kinetics.
Since all enzymes are proteins, and proteins are differentially soluble in salt solutions, enzyme extraction procedures often begin with salt (typically, ammonium sulfate) precipitation. On the simplest level, proteins can be divided into albumins and globulins on the basis of their solubility in dilute salts. Albumins are considered to be soluble, while globulins are insoluble. Solubility is relative, however, and as the salt concentration is increased, most proteins will precipitate.
Thus, if we homogenize a tissue in a solution that retains the enzyme in its soluble state, the enzyme can be subsequently separated from all insoluble proteins by centrifugation or filtration. The enzyme will be impure, since it will be in solution with many other proteins. If aliquots of a concentrated ammonium sulfate solution are added serially, individual proteins will begin to precipitate according to their solubility. By careful manipulation of the salt concentrations, we can produce fractions that contain purer solutions of enzymes, or at least are enriched for a given...
