Tubulin protomers undergo an extensive array of post-translational modifications to tailor microtubules to specific tasks. One such modification, the acetylation of lysine 40 of ?-tubulin, located in the lumen of microtubules, is associated with stable, long-living microtubule structures. MEC-17 was recently identified as the acetyltransferase that mediates this event. We have determined the crystal structure of the catalytic core of human MEC-17 in complex with its cofactor acetyl-CoA at 1.7Å resolution. The structure reveals that the MEC-17 core adopts a canonical Gcn5-related N-acetyltransferase (GNAT) fold that is decorated with extensive surface loops. An enzymatic analysis of 33 MEC-17 surface mutants identifies hot-spot residues for catalysis and substrate recognition. A large, evolutionarily conserved hydrophobic surface patch that is critical for enzymatic activity is identified, suggesting that specificity is achieved by interactions with the ?-tubulin substrate that extend outside of the modified surface loop. An analysis of MEC-17 mutants in Caenorhabditis elegans shows that enzymatic activity is dispensable for touch sensitivity.