Biotechnology, Nagoya University
○Atsuo Suzuki Akihito Fukushima Takashi Yamane

SAD(Single-wavelength Anomalous Dispersion) method is getting popular in protein crystallography. As an anomalous scatteror for SAD, a selenium atom genetically incorporated into protein is popular. But Se-SAD method requires usage of toxic selenomethionine and synchrotron X-ray. Here we cast a spotlight on transition metals as anomalous scatterors. Some of them are less toxic and suitable for SAD phasing with CuKα radiation.
We tested the first series of transition metals, Cr, Fe, Co, Ni, Cu, and Zn. They were introduced into protein crystals by the quick-soak method. Proteinase K and lysozyme were used as test proteins. Diffraction data from transition-metal derivatives were collected at 1.6A resolution using CuKα radiation. Anomalous scatterors were located by the program SHELXD and verified by anomalous difference-Fourier maps. Fe and Co were found to bind to both proteins. Using the Fe- or Co-derivative, phase calculation by SAD method and automatic model-building by ARP/wARP were easily performed. In the crystals, Fe or Co atoms bind directly to acidic side-chains, or bind to hydrophilic side-chains as octahedral aqua complex.