Structural basis of tubulin tyrosination by tubulin tyrosine ligase

TTL controls levels of tyrosinated tubulin in cultured hippocampal neurons. Shown is an image of hippocampal neurons at 5 days in vitro (DIV5) transfected with GFP-tagged E331Q mutant TTL (green) and stained for tyrosinated tubulin (tyr-tubulin; red). For the analyses of staining intensities, somas were traced of the transfected neuron (GFP filled) and a non-transfected neuron (control) in the same image. For each soma the average intensity was measured, and the ratio of transfected vs. non-transfected intensity was calculated for each image. Expression of E331Q mutant TTL in neurons did not affect the levels of tyrosinated tubulin. These results are shown in Figure 4A. Please note that for the tyrosinated tubulin staining a combination of cold methanol and paraformaldehyde was used, which is not optimal for the GFP stains in neurons.