We have investigated the microtubule-associated proteins (MAPs) of Tetrahymena pyriformis. Three polypeptides, with apparent molecular masses of 90 kDa, 60 kDa, and 37 kDa, co-sedimented with Taxol-stabilized mammalian microtubules. Partial amino acid sequencing of the 60kDa MAP revealed that it was a newly identified Tetrahymena protein. The sequence was homologous to Xenopus XMAP215, suggesting that the 60kDa MAP was a member of the XMAP215 family. The purified 60kDa MAP promoted the polymerization of mammalian tubulin in vitro. The 37kDa MAP was identical to Tetrahymena glyceraldehyde-3-phosphate dehydrogenase. The 90 kDa polypeptide was not characterized further, because of its low abundance.