If I recall correctly cytochrome oxidase, which I believe was the first protein purified with DDM, requires about 10x cmc in column buffers to keep it soluble. Check for papers from 1970's or 80's by S. Ferguson-Miller.
Cytochrme bc1 complex, on the other hand, is perfectly clear in 1 cmc and can be diluted from that into detergent-free buffer for spectroscopywithout becoming turbid, at least within an huor. My rule of thumb for solubilizing with DDM is 1 g/g protein (+ 1 cmc). If protein is say 10-20 g./l, the (+ 1 cmc) becomes completely insignificant. (Total protein, not the protein of interest. Probably depends on the amount of lipid too, but this is just a rule of thumb to start with.) eab Katarzyna Rudzka wrote:
Hi All, Has anyone had any luck purifying membrane proteins with DDM (n-dodecyl-D-maltoside) using concentration of detergent ~1-2 x CMC ? (Its CMC is very low: 0.009%). I would like to keep it as low as possible, so I don't have too much DDM around when I get to the crystallization step. I wonder If the amount of detergent sufficient for the protein extraction has to be determined experimentally for each protein or maybe there are some good rules of thumb. I appreciate your help. Thanks. Kasia Katarzyna Rudzka, Postdoctoral Fellow Department of Biophysics and Biophysical Chemistry Johns Hopkins University, School of Medicine Baltimore, Maryland 21205 USA
