[phenixbb] phenix.xtriage says that entire res range contains anomalous signal but chart says differently?

Nathaniel Echols NEchols at lbl.gov
Tue Aug 11 12:06:55 PDT 2009

On Aug 11, 2009, at 11:37 AM, sbiswas2 at ncsu.edu wrote:
> Is there a chance of backsoaking even when the protein has been  
> expressed
> as a selenomethionine derivative? I just overexpressed the protein by
> shutting down the methionine biosynthesis in E.coli and incorporated  
> SeMet
> instead of methionine.
> The protein has three methionine and is 26kDa

You could mass-spec the SeMet protein to make sure the expression  
worked, but these protocols rarely fail.   Assuming the incorporation  
of SeMet is nearly complete, you'd see a huge fluorescence peak even  
if the methionine sidechains were disordered.  Especially if the  
crystals diffract weakly and your overall signal-to-noise ratio is  
relatively low, it's not terribly surprising that you don't have much  
anomalous signal.  I've seen lots of data like this (sometimes with  
many more Met residues); unfortunately, I've never found any solution  
other than growing better crystals.  It might help to collect more  
frames to improve the signal, but this is also going to increase the  
radiation damage.

Nathaniel Echols
Lawrence Berkeley Lab
NEchols at lbl.gov

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