Dear Matt,
I use manganese for all my kinase buffers. My holobuffer contains 2mM
manganese at pH 7.0. The key was to add manganese after you pHed the
buffer with NaOH. I never used any basic buffer and it could be issue
if your buffer is too basic. Below is the recipe for my holobuffer. I
hope this helps.
Good luck,
Choel
HOLO BUFFER with ATP 2liters
MOPS 10 mM 4.2 g ATP 0.2mM 0.22g
NaCl 50mM 5.84 g TECP-HCl 1mM 0.6 g
2 mM MnCl2 0.79g pH 7.0 N2 purge
Choel Kim Ph.D.
On Apr 6, 2009, at 1:14 PM, Matthew Alan Bratkowski wrote:
Hi.
Does anyone have experience using solutions containing manganese as
crystallization buffers (buffers, not screening well solutions)? The
protein that I am working requires manganese for activity, and I
have read
reports of related proteins crystallizing in manganese buffers. I
made a
buffer containing 3 mM manganese, that initially had a black color
then
turned to deep purple, and later almost clear. After a few weeks, the
buffer turned an orange color and contains dark manganese syrup on the
bottom. Does anyone know how to prepare a manganese buffer that is
stable
enough to be used as a protein buffer for crystallization screens?
Thanks,
Matt
