Dear all,
We work with lipid droplet binding proteins in our group. These
proteins behave more like membrane associated proteins, have a very hydrophobic
surface and require detergents right from solubilization to crytallization.
We use 1% DDM for solubilization, 0.03% DDM throughout affinity
chromatography and exchange 3X CMC of OG and LDAO in gel-filtration column for
crystallization. Previously, we used to use concentrators with 30kda cut off but
recently got suggestions to use higher cut off concentrators. Our proteins arent
generally big and are less than 100kda.
Now my question is that, we havent really got any crystals over a one
year period and is it that the 3X CMC of detergents is too much for these
globular proteins in the sense it is engulfing the protein on a whole and
blocking crystal contacts? Is there a way i could play around with detergents
getting closer towards crystals?
Looking forward to valuable suggestions and thanks very much in
anticipation.
Sincerely,
Vasan
