[phenixbb] anomalous difference map

[Nathaniel Echols]

On Fri, Mar 25, 2011 at 9:20 PM, Jason <phenix.upitt at gmail.com> wrote:
> 1) Synchrotron Fluorescence scan indicates the heavy atom definitely
> presents in my protein (my control is a second protein with the same ligand
> showed no absorbance spectrum)

Keep in mind that a positive fluorescence scan does not necessarily
mean that the element of interest is bound and well-ordered in the
crystal - it is easy to get an excellent scan without seeing anything
in the maps later.

> When I load the mtz file to phenix.maps GUI, the mtz label pulldown menu
> indicates 4 possible choices for the column to use:(1) IMEAN, SIGIMEAN
> (2) I(+),  sigI(+),  I(-), sigI(-) merged (3) F(+), sigF(+), F(-), sigF(-)
> merged (4) F, sigF, Dano SigDano. I have tried the choices of (2) (3) and
> (4). However, there is barely any anomalous signal at 4sigma, which makes me
> wondering if something is not right. The first thing coming to my mind is
> the mtz labels: I(+),  sigI(+),  I(-), sigI(-) merged. What does merged
> mean? Could this be the reason? Other issues that could causing the trouble?

The "merged" means that the input data were only partially merged -
this usually happens when processing in HKL2000 using the "no merge
original index" setting, where the reflections are not merged to the
asymmetric unit (anomalous or not).  I've never used XDS, but I guess
it must do something similar.  Phenix doesn't really deal with data
like that; it always merges equivalents (while leaving Friedel pairs
alone by default).  This usually doesn't have any impact on the
anomalous signal.  If XSCALE has an option to merge the data more
completely, this should make the "merged" tag go away.

It sounds like your element of interest isn't very well ordered; do
you see the rest of the ligand in the normal maps?  If you really want
to be sure that it's not a data-handling issue, you could try
reprocessing in other programs and confirming the result.  I don't
know if radiation damage could be at fault, but it's always a
possibility.

-Nat