The ring-cleaving dioxygenases are a family of non-heme Fe(II) enzymes, of which the catechol extradiol dioxygenases are the best known. Less well known are the hydroquinone dioxygenase. The enzyme PcpA, which is structurally related to the extradiol dioxygenases, is one of the few that has been characterized. We have shown that PcpA is specific for ortho-disubstituted hydroquinones, but not catechols, with chloro- or bromo- (but not fluoro-) substituents greatly preferred at the ortho positions. Substrate binding titrations show only a small shift in pKa between the free vs. enzyme-bound substrate, suggesting that PcpA lacks an active site base needed to deprotonate the substrate (as is found in extradiol dioxygenases), thus explaining the lack of activity with catechols. We have used model complexes to examine possible interactions between halogen substituents of substrates/inhibitors and the metal center. Structurally characterized model complexes of the form (TpPh2)ML, where TpPh2 = tris(3,5- diphenyl-pyrazolyl)borate, M = Fe(II), Co(II), or Ni(II) and L = 2,6-dichloro/dibromophenolate or 2,5-dichlorohydroquinonate exhibited metal-halogen secondary bonding, with M-X distances of &lt; 2.9 Å. Variable temperature NMR studies and DFT calculations showed that the barrier to rotation of the phenolate is ~30 kJ/mol, which is on par with the strength of a hydrogen bond. Thus, metal-halogen secondary bonding could be one of several factors that govern substrate binding in the enzyme PcpA. More recently, we have explored the enzyme MnpC, which is a homolog of PcpA. This enzyme, which is believed to be an aminohydroquinone dioxygenase, exhibits hydroquinone ring-cleavage activity, but appears to differ in its substrate specificity.

Hydroquinone ring-cleaving dioxygenases: Enzymes and model complexes

254th National Meeting (2017)

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