Circular dichroism of protocatechuate 3,4-dioxygenase from Pseudomonas aeruginosa.

The circular dichroism spectrum of protocatechuate 3,4-dioxygenase from Pseudomonas aeruginosa has been examined in the absence of the substrates, protocatechuic acid and O-2 and in the presence of the competitive inhibitors, protocatechualdehyde and p-hydroxybenzoic acid. The native enzyme has a low alpha-helical content (less than 1%) and exhibits several positive ellipticity bands between 250 and 300 nm (255,269,275 and 292 nm) and two, low intensity, negative bands at 330 and 480 nm. In the presence of protocatechuic acid and the absence of O-2, spectral changes are evident in the side chain and visible regions. There is a shift in the aromatic-region maximum from 275 to 267 and in the visible region from 330 to 348 and from 480 to 555 nm. No spectral changes are observed upon the removal or addition of only O-2. Different spectral changes in both the side chain and visible regions are observed in the enzyme with the two competitive inhibitors under either aerobic or anaerobic conditions. The spectral changes observed in the side chain region suggest the possible participation of aromatic residues in the binding process, but it is not yet established as to whether these residues play an active or passive role in binding and/or catalysis.