Purpose
The protocol describes how to prepare selenomethionyl (Se-Met) protein using a regular E.coli strain. Selenium can be used for phase determination in multi-wavelength anomalous diffraction (MAD) method. Se-Met can often replace methionine residues in a protein without affecting the protein's properties, therefore producing a protein advantageous for crystal structure solving. Also, the X-ray absorption edge of selenium is easily accessible by synchrotron radiation, making a Se-Met crystal ideal for collecting anomalous X-ray diffraction data. The Se-Met proteins can also be prepared using insect cells and CHO cells, which will be described in separate protocols.
Materials
LB media
antibiotics (1000x conc.)
1M IPTG
M9 media (minimal media)
1 Liter 5x M9 media: (sterile filtered)