Comparative analysis of catalases: spectral evidence against heme-bound water for the solution enzymes.

A recent X-ray structural analysis of M. luteus catalase indicates heme-bound H2O trans to the proximal tyrosinate ligand, a finding in contrast to previous X-ray data reporting a 5-coordinate heme for bovine liver catalase. The presence of heme-bound H2O, requiring displacement prior to substrate-binding, is likely to be catalytically significant for catalases. We have used magnetic circular dichroism (MCD) spectroscopy, a highly accurate method for assignment of heme spin- and coordination-states, to study native, solution forms of bovine liver, M. luteus, and A. niger catalases. All three enzymes display similar spectral features with the weak (approximately 5 delta epsilon M [moles.cm.Tesla]-1) intensity typical of a 5-coordinate high-spin ferric heme. No evidence for H2O-ligation, inducing a 6-coordinate heme, occurred upon variation of pH or buffer composition. Therefore, we suggest that the catalytically significant structure of catalases has an unoccupied heme binding site trans to the proximal tyrosinate heme ligand.