Affinity chromatography in nonionic detergent solutions.
Anionic dye affinity chromatography is commonly unproductive in the presence of nonionic detergents used to extract particulate proteins. Using lactate dehydrogenase as a model protein, Cibacron blue F3GA as a model dye, and Triton X-100 as a model detergent, we find that the dye is encapsulated in nonionic detergent micelles, rendering the dye incapable of ligation with the enzyme. However, the dye can be liberatd from the micelles without altering the nonionic detergent concentration by addition of an anionic detergent, such as deoxycholate or sodium dodecyl sulfate, forming mixed anionic/nonionic micelles that displace the anionic dye. Encapsulation of the anionic detergents prevents their activity as protein denaturants. These observations have been successfuly translated to the dye affinity chromatography of a detergent extract of brain particulate cyclic nucleotide phosphodiesterase.
D000818:Animals; D000873:Anthracenes; D001923:Brain Chemistry; D002417:Cattle; D002846:Chromatography, Affinity / Q000379:methods*; D004396:Coloring Agents; D003902:Detergents; D007770:L-Lactate Dehydrogenase / Q000302:isolation & purification; D008565:Membrane Proteins / Q000302:isolation & purification*; D008566:Membranes / Q000201:enzymology*; D009132:Muscles / Q000201:enzymology; D011817:Rabbits; D013057:Spectrum Analysis; D013501:Surface-Active Agents; D014227:Triazines
D000873:Anthracenes; D004396:Coloring Agents; D003902:Detergents; D008565:Membrane Proteins; D013501:Surface-Active Agents; D014227:Triazines; C012842:Cibacron Blue F 3GA; D007770:L-Lactate Dehydrogenase