Effect of Metals in Biomimetic Dimetal Complexes on Affinity and Gas-Phase Protection of Phosphate Esters

Although the biomimetic dimetal complex [LGa2(OH)2(H2O)2]3+ [L = 2,6-bis­((N,N′-bis­(2-picolyl)­amino)­methyl)-4-tertbutylphenolate] provides efficient protection against phosphate loss in phosphopeptides upon collision-induced dissociation tandem mass spectrometry (CID MS/MS), the underlying mechanism remains unknown. Here, we explored the mechanism in detail and investigated the selective binding to phosphate groups in solution. Dimetal complexes containing combinations of Ga3+, In3+, Fe3+, Co3+, Zn2+, Cu2+, and V2+ were reacted with HPO42–, phosphoserine, and a phosphopeptide (FQ­pS­EEQ­QQT­EDE­LQ­DK, abbreviated “βcas”) and studied with isothermal titration calorimetry (ITC), CID MS/MS, and density functional theory (DFT). Ka for HPO42– binding scaled with the metal charge and was 35-fold larger for [LGa2(OH)2(H2O)2]3+ (3.08 ± 0.31 × 106 M–1) than for [LZn2(HCOO)2]+. CID MS/MS of [LGa2(βcas)]n+ revealed protection against phosphate detachment (<3% of the total ion intensity). Phosphate detachment from βcas was 22–40% and increased to 42–71% when bound to dimetal complexes of lower charge than {LGa2}5+. CID data suggests that facile metal–phosphate dissociation is associated with proton transfer from the intermediate oxazoline ring formed in the phosphopeptide to the metal–phosphate complex. The observed phosphate stabilization was attributed to a significant reduction in the gas-phase basicity (GB) of the phosphate group when bound to {LGa2}5+/{LIn2}5+ complex cores. Absence of proton transfer results in formation of an ion–zwitterion intermediate with a greater dissociation threshold. This hypothesis is supported by DFT calculations for [LGa2(PO4)]2+, [LGaZn­(PO4)]+, [LZn2(PO4)], and 2,4-dimethyl-3-oxazoline showing that [LGa2(PO4)]2+ is the only compound with a substantial lower GB (321 kJ/mol less) than 2,4-dimethyl-3-oxazoline.