Dear Martin, You could calculate an Fo-Fc map with the FAD having half occupancy. This should bring out the "pure" difference density for your modified FAD, which might be easier to interpret. You may have to try different occupancies, say 0.4, 0.45, 0.5, 0.55, 0.6 etc. to find the point with the least residual intact FAD density.
In the absence of any pictures of the electron density and any information about which ring is almost missing and how the other rings are distorted, it is very difficult, if not impossible to provide you with any clues. Best, Herman Von: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] Im Auftrag von Martin Malý Gesendet: Dienstag, 7. November 2017 17:00 An: CCP4BB@JISCMAIL.AC.UK Betreff: [EXTERNAL] Re: [ccp4bb] Radiation damage to the FAD in enzyme structure Dear colleagues, thanks for all your responses. Just to make things clear, we are absolutely sure about 100% presence of the FAD in the original solution. What we see in the electron density is roughly half occupancy for the FAD and additional difference density for something what looks like a product of some FAD degradation. Either caused by degradation or age. The product looks like one of the three planar ring is almost missing, the other two are significantly distorted. We see it clearly, but we want to honestly and correctly interpret the density based on some prior chemical knowledge. Does anyone has a clue? In the mean time, we are reading the other literature provided by you, of course. Best regards, Martin ________________________________ Od: CCP4 bulletin board <CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK>> za uživatele Martin Malý <malym...@fjfi.cvut.cz<mailto:malym...@fjfi.cvut.cz>> Odesláno: 6. listopadu 2017 12:01:43 Komu: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> Předmět: [ccp4bb] Radiation damage to the FAD in enzyme structure Dear colleagues, I am investigating a structure of a FAD-dependent enzyme. The electron density map suggests radiation damage to the FAD. It apparently is different from simple change of the redox state and "butterfly"-like structure. We did not find in literature possible products of radiation damage, like a removal of several atoms of the FAD. Has anyone observed such effect? To describe it in more detail, I can observe negative difference map of C2, N3, C4, and O4 atoms of flavine. Moreover, there is positive difference map close to O2 and O4 atoms thus it looks as water molecules are bound there instead of the missing FAD atoms. I am attaching parameters of the experiment: performed at synchrotron, exposition time 210 s, high resolution diffraction limit 1.65 A (<I/sigma> = 2 at shell 1.75-1.65 A). We could see a decrease of diffraction data statistics during the experiment hence we think there is significant radiation damage to the crystal. Thank you very much for ideas. Regards, Martin Maly
