Tuning caged calcium: Photolabile analogues of EGTA with improved optical and chelation properties

Abstract

The physico-chemical properties of several Ca 2+-selective, photolabile chelators are described. These molecules have been developed as part of an effort to produce a caged Ca 2+ that improved upon the Ca 2+ chelation properties and light absorption capability of nitrophenyl-EGTA (NP-EGTA). Four dimethoxy- ortho-nitrophenyl derivatives of EGTA (called DMNPE-1 through -4), and one analogue of EGTA (DMNPE-5) have been characterized, each of which is bisected upon irradiation. One of these cages has a higher affinity than NP-EGTA: DMNPE-4 has a K d for Ca 2+ of 48 nm at pH 7.2 (19 nM at pH 7.4). Furthermore, this cage has a large extinction coefficient of 5120 M −1 cm −1 at 350 nm (cf. 975 M −1 cm −1 for NP-EGTA). The other physico-chemical properties of DMNPE-4 are: quantum yield of photolysis of 0.09; bipasic Ca 2+ release kinetics (70% released with a rate of about 48,000 s −1 and 30% at 1.5 s −1) and photoproducts that bind Ca 2+ with very low affinity ( K d in the range of 2 mM, pH 7.2), hence most of the bound Ca 2+ is released rapidly and efficiently upon photolysis. Thus, DMNPE-4 has a unique combination of properties that make it an extremely effective Ca 2+ cage.