We used cross-linking mass spectrometry with the cleavable cross-linker, disuccinimidyl dibutyric urea (DSBU) to map sites of interaction between the proline rich domain (PRR, tau amino acids 149-244) and tubulin. For comparison, we also conducted these experiments with PRR constructs containing multiple phosphomimic mutations (from serine or threonine to glutamate) at tau amino acid positions 181, 217, 231 and 235.