Binding of NO and CO to the d(1) heme of cd(1) nitrite reductase from Pseudomonas aeruginosa

The cd(1) nitrite reductase, a key enzyme in bacterial denitrification, cat alyzes the one-electron reduction of nitrite to nitric oxide. The enzyme co ntains two redox centers, a c-type heme and a unique d(1) heme, which is a dioxoisobacteriochlorin. Nitric oxide, generated by this enzymatic pathway, if not removed from the medium, can bind to the ferrous d(1) cofactor with extremely high affinity and inhibit enzyme activity. In this paper, we rep ort the resonance Raman investigation of the properties of nitric oxide and carbon monoxide binding to the d(1) site of the reduced enzyme. The Fe-lig and (Fe-NO and Fe-CO) stretching vibrational frequencies are unusually high in comparison to those of other ferrous heme complexes. The frequencies of the Fe-NO and N-O stretching modes appear at 585 and 1626 cm respectively, in the NO complex, while the frequencies of the Fe-CO and C-O stretching m odes are at 563 and 1972 cm(-1), respectively, for the CO complex. Also, th e widths (fwhm) of the Fe-CO and C-O stretching modes are smaller than thos e observed in the corresponding complexes of other heme proteins. The unusu al spectroscopic characteristics of the d(1) cofactor are discussed in term s of both its unique electronic properties and the strongly polar distal en vironment around the iron-bound ligand. It is likely that the influence of a highly ruffled structure of heme d(1) on its electronic properties is the major factor causing, anomalous Fe-ligand vibrational frequencies.