Isolation of a Membrane-Bound Enzyme, 5-Nucleotidase

Membrane enzymes may be peripheral or integral proteins. The attachment of peripheral enzymes is mediated mainly by association with other membrane proteins and consists primarily of hydrophilic, electrostatic interactions. Integral membrane proteins are primarily bound by hydrophobic interactions with the lipid bilayer, either via one or more transmembrane peptide domains (1 ) or by possession of a glycosyl-phosphatidylinositol (GPI) membrane anchor at the carboxy-terminus (2 ). The purification of a membrane enzyme thus requires two additional considerations compared to the isolation of a cytosolic enzyme: the isolation of a membrane fraction containing the enzyme and the solubilization of the enzyme from this fraction. Once solubilization has been achieved and maintained, the enzyme can be treated essentially as a soluble enzyme and subjected to a wide range of purification techniques.