Sequential assignment of proline-rich regions in proteins: Application to modular binding domain complexes

Many protein-protein interactions involve amino acid sequences containing proline-rich motifs and even poly-proline stretches. The lack of amide protons in such regions complicates assignment, since (HN)-H-1-based triple-resonance assignment strategies cannot be employed. Two such systems that we are currently studying include an SH2 domain from the protein Crk with a region containing 9 prolines in a 14 amino acid sequence, as well as a WW domain that interacts with a proline-rich target. A modified version of the HACAN pulse scheme, originally described by Bax and co-workers [Wang et al. (1995) J. Biomol. NMR, 5, 376-382], and an experiment which correlates the intra-residue H-1(alpha), C-13(alpha)/C-13(beta) chemical shifts with the N-15 shift of the subsequent residue are presented and applied to the two systems listed above, allowing sequential assignment of the molecules.