Multiple heavy-atom reagents for macromolecular X-ray structure determination. Application to the nucleosome core particle.

The X-ray structure of the nucleosome core particle was solved at 7 A resolution using the method of multiple isomorphous replacement based on two isomorphous derivatives, each containing a different multiple heavy-atom compound. The preparation of these heavy-atom compounds and their application to this macromolecular structure determination are described. The first of these reagents, TAMM (tetrakis(acetoxymercuri)methane), was solubilized by the addition of an excess of glycylglycine and, when added to crystals of the nucleosome core particle, produced a derivative with a single major site. Despite the large mass of 206,000 daltons per asymmetric unit, the position of the TAMM molecule was found in these crystals using the difference Patterson technique. This compound was sufficiently electron-dense to produce a unique solution, whereas the mono-mercurial, methylmercury nitrate had been inadequate. The second reagent, PIP (di-mu-iodobis(ethylenediamine)diplatinum(II) nitrate), is freely soluble in aqueous solution and, on addition to the crystals, labelled the histone proteins at several sites. The locations of the PIP groups were determined from difference Fourier and Patterson maps. The X-ray structure and solution characterization of this compound are reported. These multiple heavy-atom compounds appear to be generally applicable to X-ray structure determination, and are particularly useful in conjunction with crystals having asymmetric units of large volume but lacking non-crystallographic symmetry elements.