Fig. 1. The analogue-sensitive Don3^M157A kinase is active in vivo. (A) Alignment of the ATP-binding pocket of the U. maydis (Um) Don3 kinase domain with corresponding sequences of other yeast (Sc) and human (Hs) kinases, which have been used to create analogue-sensitive kinases. The gatekeeper amino acids (positions 140 and 157) are bold and the position of the amino acid exchange is indicated. (B) Pcrg::don3 and Pcrg::don3^M157G/A expression cassettes. The kinase domain (KD) is indicated, and the site of the amino acid exchange is marked by an asterisk. (C) don3 cells expressing Don3, Don3^M157G and Don3^M157A under control of the inducible Pcrg-promoter. DIC-images illustrate the repressed state (glucose) and the induced state (arabinose). Scale bars, 10 µm.