Abstract
The dissimilatory nitrite reductase (cytochromec,dl) from Pseudomonas aeruginosa was observed at pH 7.5 to catalyzenitrosytlransfer (nitrosation) between [I6N]nitriteand several N-nucleophiles orH2180,with rate enhancement of the order of lo8relative toanalogous chemical reactions. Thereducing system (ascorbate, N,N,N',N'-tetramethylphenylenediamine)could reduce nitrite (but not NO) enzymatically and had essentially no direct chemical reactivity toward nitrite or NO. The N-nitrosations showed saturation kinetics with respect to the nucleophile and, while exhibiting V, values which varied by about 40-fold, nevertheless showed little or no dependence of V, on nucleophile pK,. The N-nitrosations and NO;/H20-'80 ex- change required the reducing system, whereas NO/ HzO-"O exchange was inhibited by the reducing system. NO was not detected to serve as a nitrosyl donor to N-nucleophiles. These and other kinetic observations suggest that the enzymatic nitrosyl donor is an enzyme-bound species derived from reduced enzyme and one molecule of nitrite, possibly a heme-nitrosyl compound (E-Fe".NO+) for which thereis precedence. Nitrosyl transfer to N-nucleophiles may occur within a ternary complex of enzyme, nitrite,and nucleophile. Catalysis of nitrosyl transfer by nitrite reductase represents a new class of enzymatic reactions and may present another example of electrophilic catalysis by a metal center. The nitrosyl donor trapped by these reactions is believed to represent an intermediate in the reduction of nitriteby cytochrome c,dl.