SelenoMet-based structure

Chris Larsen clarsen at
Fri Sep 23 07:48:39 EST 1994


I have a protein that my collaborator has been trying to crystallize for 
three years now.  We have a native data set to 3.8 A. Unfortunately,
the heavy atom derivative has been unstable.

My strain is the BL21 variety, and uses a T7 promoter.  I think that the 
met auxotroph of this strain is available, so that a seleno met version 
could easily be created.  I can purify 100 mg of this 25 kDa protein to
a single band on an overloaded silver-stained gel in ten days.
The crystallization conditions for this orthorhombic P2P1P1 crystal is 
defined for the native protein. 

My questions are:

Is the anomolous scattering (?) method using a synchrotron 
dependent on the native data set already obtained?

Can all the data be collected on a single crystal? Is a second derivative necessary?

Would this process be more difficult than the standard methods?

Does the difficulty depend on the number of Met in the protein?

AND:  Is it feasible to do?

Thanks heaps......


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