TY - JOUR
T1 - Radiation-induced site-specific damage of mercury derivatives
T2 - Phasing and implications
AU - Ramagopal, Udupi A.
AU - Dauter, Zbigniew
AU - Thirumuruhan, Radhakannan
AU - Fedorov, Elena
AU - Almo, Steven C.
PY - 2005/9
Y1 - 2005/9
N2 - The behavior of mercury-derivatized triclinic crystals of a 60 kDa protein target from the New York Structural GenomiX Research Consortium provides novel insights into the mechanism of heavy-atom-specific radiation damage and its potential exploitation for de novo structure solution. Despite significant anomalous signal, structure solution by classic SAD and MAD phasing approaches was not successful. A detailed analysis revealed that significant isomorphic variation of the diffracted intensities was induced by X-ray irradiation. These intensity changes allowed the crystal structure to be solved by the radiation-damage-induced phasing (RIP) technique. Inspection of the crystal structure and electron-density maps demonstrated that the covalent S-Hg bonds at all four derivatized cysteine sites were much more susceptible to radiation-induced cleavage than other bonds typically present in native proteins. A simple diagnostic is described to identify the fingerprint of such decay at the time of data collection/processing. The rapid radiation-induced decomposition of mercury adducts is consistent with the difficulties frequently associated with the experimental phasing of mercury derivatives and suggests a straightforward solution to overcome this limitation by radiation-damage-induced phasing with anomalous scattering (RIPAS). These results indicate that historically recalcitrant and newly emerging difficulties associated with Hg phasing should be revisited.
AB - The behavior of mercury-derivatized triclinic crystals of a 60 kDa protein target from the New York Structural GenomiX Research Consortium provides novel insights into the mechanism of heavy-atom-specific radiation damage and its potential exploitation for de novo structure solution. Despite significant anomalous signal, structure solution by classic SAD and MAD phasing approaches was not successful. A detailed analysis revealed that significant isomorphic variation of the diffracted intensities was induced by X-ray irradiation. These intensity changes allowed the crystal structure to be solved by the radiation-damage-induced phasing (RIP) technique. Inspection of the crystal structure and electron-density maps demonstrated that the covalent S-Hg bonds at all four derivatized cysteine sites were much more susceptible to radiation-induced cleavage than other bonds typically present in native proteins. A simple diagnostic is described to identify the fingerprint of such decay at the time of data collection/processing. The rapid radiation-induced decomposition of mercury adducts is consistent with the difficulties frequently associated with the experimental phasing of mercury derivatives and suggests a straightforward solution to overcome this limitation by radiation-damage-induced phasing with anomalous scattering (RIPAS). These results indicate that historically recalcitrant and newly emerging difficulties associated with Hg phasing should be revisited.
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U2 - 10.1107/S0907444905022316
DO - 10.1107/S0907444905022316
M3 - Article
C2 - 16131763
AN - SCOPUS:32944455892
SN - 0907-4449
VL - 61
SP - 1289
EP - 1298
JO - Acta Crystallographica Section D: Biological Crystallography
JF - Acta Crystallographica Section D: Biological Crystallography
IS - 9
ER -