| Popis: |
Membrane proteins tend to form aggregates even in the presence of detergents, which reduces the efficacy of all separation techniques. It is therefore essential to select an effective solubilizing, but non-denaturing detergent to ascertain the minimum ionic strength the protein requires to be in a homogeneously dissolved state, the maximum ionic strength that will not cause dissociation of labile proteins from complex structures, and the optimum pH for solubilization and protein stability. In planning a strategy for membrane protein isolation, it is necessary to use methods that preserve the native state of the protein in the first steps of the purification. If enzymatic activity is not required, application of denaturing techniques may be advantageous in the final stages when proteins can be identified by apparent molecular masses on SDS-PAGE. Enriching the protein of interest before adding detergents facilitates purification. If there is a choice, attempts should be made to isolate the protein from sources that contain high levels of it. |
