TY - JOUR
T1 - Time-dependent analysis of K2PtBr6 binding to lysozyme studied by protein powder and single crystal X-ray analysis
AU - Helliwell, John R.
AU - Tony Bell, A. M.
AU - Bryant, Pat
AU - Fisher, Stu J.
AU - Habash, George
AU - Helliwell, Madeleine
AU - Margiolaki, Irene
AU - Kaenket, Surasak
AU - Watier, Yves
AU - Wright, Jon
AU - Yalamanchilli, Sampath
PY - 2010
Y1 - 2010
N2 - Multi heavy atom cluster compounds like K2PtBr6 offer a way forward to solve, de novo, unknown protein structures by powder diffraction involving dispersive (measured at two X-ray wavelengths) and isomorphous intensity differences as a complement to micro-crystallography or by using both approaches in combination. Towards this end, using the ESRF high resolution synchrotron X-ray powder diffraction beamline ID31, we have recorded high quality protein powder diffractograms at the platinum LIII and bromine K absorption edges, as well as reference wavelengths, for K2PtBr6 bound to lysozyme. These experiments were conducted at 80K to protect the sample against X-radiation damage as much as possible and also to fix the K2PtBr6 bound state, which seemed to show instability at room temperature. With multiple powder pattern analysis we extracted intensities and showed the PtBr6 bound in lysozyme using 'omit electron density maps'. In addition the wavelength dispersive Fourier around the Br K edge shows up the bromine signal at PtBr6 binding site 1 in one of the six samples tested. To better understand the chemical properties of this heavy atom compound we have elucidated the detailed binding behaviour using single crystal analyses with time-resolved freeze quenching after soak times of 10, 90 and 170 minutes. Whilst the quick soaking of 10 to 30 minutes, used at ESRF ID31 shows clear binding, there is increasing binding strength with increasing soak time. Thus, these time-resolved analytical chemistry results show that further heavy atom signal optimizations are possible. Prospects for extending our approach to the yet larger isomorphous and wavelength dispersive signal case of Ta6Br12 bound to lysozyme are also described.
AB - Multi heavy atom cluster compounds like K2PtBr6 offer a way forward to solve, de novo, unknown protein structures by powder diffraction involving dispersive (measured at two X-ray wavelengths) and isomorphous intensity differences as a complement to micro-crystallography or by using both approaches in combination. Towards this end, using the ESRF high resolution synchrotron X-ray powder diffraction beamline ID31, we have recorded high quality protein powder diffractograms at the platinum LIII and bromine K absorption edges, as well as reference wavelengths, for K2PtBr6 bound to lysozyme. These experiments were conducted at 80K to protect the sample against X-radiation damage as much as possible and also to fix the K2PtBr6 bound state, which seemed to show instability at room temperature. With multiple powder pattern analysis we extracted intensities and showed the PtBr6 bound in lysozyme using 'omit electron density maps'. In addition the wavelength dispersive Fourier around the Br K edge shows up the bromine signal at PtBr6 binding site 1 in one of the six samples tested. To better understand the chemical properties of this heavy atom compound we have elucidated the detailed binding behaviour using single crystal analyses with time-resolved freeze quenching after soak times of 10, 90 and 170 minutes. Whilst the quick soaking of 10 to 30 minutes, used at ESRF ID31 shows clear binding, there is increasing binding strength with increasing soak time. Thus, these time-resolved analytical chemistry results show that further heavy atom signal optimizations are possible. Prospects for extending our approach to the yet larger isomorphous and wavelength dispersive signal case of Ta6Br12 bound to lysozyme are also described.
U2 - 10.1524/zkri.2010.1349
DO - 10.1524/zkri.2010.1349
M3 - Article
SN - 0044-2968
VL - 225
SP - 570
EP - 575
JO - Zeitschrift fur Kristallographie
JF - Zeitschrift fur Kristallographie
IS - 12
ER -