title: Mapping protein-ligand interactions in the gas phase using a functional group replacement strategy. Comparison of CID and BIRD activation methods.
abstract: Intermolecular interactions in the gaseous ions of two protein-ligand complexes, a single chain antibody (scFv) and its trisaccharide ligand (α-D-Galp-(1→2)-[α-D-Abep-(1→3)]-α-Manp-OCH3, L1) and streptavidin homotetramer (S4) 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 (S4 + 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 (Ec50) 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 Ec50 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 (S4 + 4B)(13+) ion. In fact, mutation of Trp79 and Trp120 led to an increase in the Ec50 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.
authors.affiliation: Department of Chemistry and Alberta Glycomics Centre, University of Alberta, Edmonton, Alberta, Canada, T6G 2G2.
authors.forename: Lu
authors.initials: L
authors.lastname: Deng
authors.forename: Elena N
authors.initials: EN
authors.lastname: Kitova
authors.forename: John S
authors.initials: JS
authors.lastname: Klassen
mesh_terms: D000818:Animals; D002412:Cations / Q000737:chemistry; D005740:Gases; D006860:Hydrogen Bonding; D007259:Infrared Rays; D008024:Ligands; D025941:Protein Interaction Mapping / Q000379:methods*; D057127:Single-Chain Antibodies / Q000737:chemistry; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D019809:Streptavidin / Q000737:chemistry; D014312:Trisaccharides / Q000737:chemistry
chemical_list: D002412:Cations; D005740:Gases; D008024:Ligands; D057127:Single-Chain Antibodies; D014312:Trisaccharides; D019809:Streptavidin