Membrane and Particulate-Associated Proteins
Membrane proteins are chemically much the same as soluble proteins, with a similar amino acid profile, although they do contain, on average, a slightly higher proportion of nonpolar amino acids. Following cell disruption in a neutral, isotonic buffer that does not contain detergents, many proteins are associated with insoluble components of the cell extract. Integral membrane proteins, material trapped within an organelle, and other proteins that are strongly associated with the cytoskeletal matrix will all be found in the particulate fraction. When one considers that the cytosol is responsible for roughly 75% of cellular protein, a protein exclusively found in the precipitate will be enriched fourfold over the crude extract. This section discusses strategies for dissociating a membrane into its individual components in such a way that the protein of interest retains its bioactivity and becomes amenable to standard purification and analytical procedures. Since there exists no single procedure to characterize the different types of membrane proteins, several methods will be presented to begin this process.
KeywordsMembrane Protein Critical Micelle Concentration Integral Membrane Protein Sodium Cholate Detergent Concentration
Unable to display preview. Download preview PDF.
- Bennett JP (1982): Solubilisation of membrane-bound enzymes and analysis of membrane protein concentration. In: Techniques in Lipid and Membrane Biochemistry. Holland: ElsevierGoogle Scholar
- Bordier C (1981): Phase separation of integral membrane proteins in Triton X-114. J Biol Chem 256: 1604–1607Google Scholar
- Creighton TE (1984): Proteins: Structures and Molecular Properties. New York: WH FreemanGoogle Scholar
- King TE (1963): Reconstitution of respiratory chain enzyme systems. J Biol Chem 238: 4037–4051Google Scholar
- Nelson N, Eytan E, Notsani B-E, Sigrist H, Sigrist-Nelson K, Gitler C (1977): Isolation of a chloroplast N,N’-dicyclohexylcarbodiimide-binding proteolipid, active in proton translocation. Proc Natl Acad Sci USA 74: 2375–2378Google Scholar
- Okumura T, Hasitz M, Jamieson GA (1978): Platelet glycocalicin. Interaction with thrombin and role as thrombin receptor of the platelet surface. J Biol Chem 253: 3435–3443Google Scholar
- Pierce Catalog and Handbook: Life Science and Analytical Research Products (1994): Rockford. IL: Pierce Co.Google Scholar
- Rivnay B, Metzger H (1982): Reconstitution of the receptor for immunoglobulin E into liposomes. J Biol Chem 257: 12800–12808Google Scholar
- Tanner MJA, Boxer DH (1972): Separation and properties of the major proteins of the human erythrocyte membrane. Biochem J 129: 333–347Google Scholar