Distinct structural and redox properties of the heme active site in bacterial dye decolorizing peroxidase-type peroxidases from two subfamilies: resonance Raman and electrochemical study.

Spectroscopic data of dye decolorizing peroxidases (DyPs) from Bacillus subtilis (BsDyP), an A subfamily member, and Pseudomonas putida (PpDyP), a B subfamily enzyme, reveal distinct heme coordination patterns of the respective active sites. In solution, both enzymes show a heterogeneous spin population, with the six-coordinated low-spin state being the most populated in the former and the five-coordinated quantum mechanically mixed-spin state in the latter. We ascribe the poor catalytic activity of BsDyP to the presence of a catalytically incompetent six-coordinated low-spin population. The spin populations of the two DyPs are sensitively dependent on the pH, temperature, and physical, i.e., solution versus crystal versus immobilized, state of the enzymes. We observe a redox potential for the Fe(2+)/Fe(3+) couple in BsDyP (-40 mV) at pH 7.6 substantially more positive than those reported for the majority of other peroxidases, including PpDyP (-260 mV). Furthermore, we evaluate the potential of the studied enzymes for biotechnological applications on the basis of electrochemical and spectroelectrochemical data.