Spectroscopic properties of the states of pig pancreatic phospholipase A2 at interfaces and their possible molecular origin.

The near-UV absorption and fluorescence spectroscopic properties of Trp-3 of pig pancreatic phospholipase A2 (PLA2) in aqueous solution (E form) or at the interface without (E* form) or with a ligand at the active site (E*L form) are characterized. In the E form, the single tryptophan residue is exposed on the protein surface to the aqueous environment, as it is freely accessible to aqueous quenchers such as succinimide and acrylamide. The fluorescence quantum yield of E is about one-third that of N-acetyl-tryptophanamide, indicating significant intramolecular quenching processes including charge-transfer reactions, as seen by the D2O effect. Upon binding of PLA2 to micelles of 1-hexadecylpropanediol-3-phosphocholine (E*), a positive difference spectrum with a shoulder at 284 nm (delta epsilon = 370 M-1 cm-1) is observed. Similar difference spectra are also observed upon binding of sulfate ion to the E form. The fluorescence emission of E* is blue-shifted by about 10 nm to 336 nm, with a 2-fold higher quantum yield. Trp-3 in E* is significantly shielded from aqueous quenchers, and the D2O effect on the quantum yield is still present. The UV difference spectrum for the E*-to-E*L transition is of large amplitude with peaks at 292 (delta epsilon = 2540 M-1 cm-1) and 284 nm (delta epsilon = 2100 M-1 cm-1), which suggests transfer of tryptophan from an aqueous to a less polar environment. Upon conversion to the E*L form, there is a further blue shift to 333 nm, with about a 20% increase in the fluorescence quantum yield.(ABSTRACT TRUNCATED AT 250 WORDS)