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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