Mapping Protein-Ligand Interactions in the Gas Phase Using a Functional Group Replacement Strategy. Comparison of CID and BIRD Activation Methods

Intermolecular interactions in the gaseous ions of two protein-ligand complexes, a single chain antibody (scFv) and its trisaccharide ligand (alpha-D-Galp-(1 -> 2)-[alpha-D-Abep-(1 -> 3)]-alpha-Manp-OCH3, L1) and streptavidin homotetramer (S-4) and biotin (B), were investigated using a collision-induced dissociation (CID)-functional group replacement (FGR) strategy. CID was performed on protonated ions of a series of structurally related complexes based on the (scFv + L1) and (S-4 + 4B) complexes, at the +10 and +13 charge states, respectively. Intermolecular interactions were identified from decreases in the collision energy required to dissociate 50 % of the reactant ion (Ec(50)) upon modification of protein residues or ligand functional groups. For the (scFv + L1)(10+) ion, it was found that deoxygenation of L1 (at Gal C3 and C6 and Man C4 and C6) or mutation of His101 (to Ala) resulted in a decrease in Ec(50) values. These results suggest that the four hydroxyl groups and His101 participate in intermolecular H-bonds. These findings agree with those obtained using the blackbody infrared radiative dissociation (BIRD)-FGR method. However, the CID-FGR method failed to reveal the relative strengths of the intermolecular interactions or establish Man C4 OH and His101 as an H-bond donor/acceptor pair. The CID-FGR method correctly identified Tyr43, but not Ser27, Trp79, and Trp120, as a stabilizing contact in the (S-4 + 4B)(13+) ion. In fact, mutation of Trp79 and Trp120 led to an increase in the Ec(50) value. Taken together, these results suggest that the CID-FGR method, as implemented here, does not represent a reliable approach for identifying interactions in the gaseous protein-ligand complexes.