In order to understand heme environment of a heme-regulated phosphodiesterase (Ec DOS), the binding behavior of cyanide to the Fe (III) complex was examined. Interestingly, the rate of cyanide binding to full-length Ec DOS was unusually slow with k(on) = 0.0022 mM(-1) s(-1), while the rate for the isolated heme domain of Ec DOS (0.045 mM(-1) s(-1)) was 20-fold higher. Ala and Leu mutations at Met95, which has been suggested to be a heme axial ligand. increased the k(on) rate 11 - and 8-fold, respectively, and dramatically decreased the cyanide dissociation rate from the isolated heme domain. His mutation at Met95, on the other hand, caused a 17-fold decrease in the k(on) value. We discuss the unusual cyanide binding behavior and the role of Met95 in controlling cyanide binding. (C) 2002 Elsevier Science (USA). All rights reserved.