High-Resolution Cryo-Electron Microscopy of Two-Dimensional Protein Crystals

The task in investigating the structure of crystals is to measure the Fourier coefficients in amplitude and phase. Electron crystallography, i.e. electron diffraction combined with electron-microscopical imaging, is a straigthforward method, because there is a priori no “phase problem”. But radiation damage during electron exposure was for a long time an absolutely unbridgeable chasm on the way to high-resolution structure research of two-dimensional protein crystals by electron microscopy. Now, using cryo-electron microscopy and computer image processing, it has been proven possible to overcome this difficulty and achieve high resolution.Here we present a rough description of the recording procedure. We used a helium-cooled superconducting objective lens with 4.5 K specimen temperature, but the procedure should also be applicable at higher specimen temperatures. The radiation damage is reduced at low specimen temperature, but unfortunately there still remains a remarkable deterioration of the crystal with increasing dose, which means that minimum exposure is advisable.