New methods to prepare iodinated derivatives by vaporizing iodine labelling (VIL) and hydrogen peroxide VIL (HYPER-VIL)

New techniques are presented for the preparation of iodine derivatives, involving vapour diffusion of iodine. Firstly, in the vaporizing iodine labelling (VIL) technique, a small amount of KI/I₂ solution is enclosed in a crystallization well, with the result that gaseous I₂ molecules diffuse into the crystallization droplets without exerting substantial changes in ionic strength in the target crystals. Once they have diffused into the droplet, the I₂ molecules sometimes iodinate accessible tyrosines at ortho positions. Secondly, when iodination is insufficient, the hydrogen peroxide VIL (HYPER-VIL) technique can be further applied to increase the iodination ratio by the addition of a small droplet of hydrogen peroxide (H₂O₂) to the crystallization well; the gaseous H₂O₂ also diffuses into the crystallization droplet to emphasize the iodination. These techniques are most effective for phase determination when coupled with softer X-rays, such as those from Cu Kα or Cr Kα radiation. The effectiveness of these techniques was assessed using five different crystals. Four of the crystals were successfully iodinated, providing sufficient phasing power for structure determination.