Observation of fast release of NO from ferrous d₁ haem allows formulation of a unified reaction mechanism for cytochrome cd₁ nitrite reductases.
Cytochrome cd1 nitrite reductase is a haem-containing enzyme responsible for the reduction of nitrite into NO, a key step in the anaerobic respiratory process of denitrification. The active site of cytochrome cd1 contains the unique d1 haem cofactor, from which NO must be released. In general, reduced haems bind NO tightly relative to oxidized haems. In the present paper, we present experimental evidence that the reduced d1 haem of cytochrome cd1 from Paracoccus pantotrophus releases NO rapidly (k=65-200 s(-1)); this result suggests that NO release is the rate-limiting step of the catalytic cycle (turnover number=72 s(-1)). We also demonstrate, using a complex of the d1 haem and apomyoglobin, that the rapid dissociation of NO is largely controlled by the d1 haem cofactor itself. We present a reaction mechanism proposed to be applicable to all cytochromes cd1 and conclude that the d1 haem has evolved to have low affinity for NO, as compared with other ferrous haems.
Dipartimento di Scienze Biochimiche A. Rossi Fanelli, Sapienza - Università di Roma, P A Moro, 5 00185 Rome, Italy.
Serena
S
Rinaldo
Katharine A
KA
Sam
Nicoletta
N
Castiglione
Valentina
V
Stelitano
Alessandro
A
Arcovito
Maurizio
M
Brunori
James W A
JW
Allen
Stuart J
SJ
Ferguson
Francesca
F
Cutruzzolà
D001059:Apoproteins / Q000378:metabolism; D001426:Bacterial Proteins / Q000378:metabolism*; D055162:Biocatalysis; D003580:Cytochromes / Q000378:metabolism*; D058440:Denitrification; D006418:Heme / Q000031:analogs & derivatives* / Q000378:metabolism; D007700:Kinetics; D008958:Models, Molecular; D009211:Myoglobin / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism*; D009572:Nitrite Reductases / Q000378:metabolism*; D010084:Oxidation-Reduction; D042084:Paracoccus pantotrophus / Q000201:enzymology*; D010782:Photolysis
D001059:Apoproteins; D001426:Bacterial Proteins; D003580:Cytochromes; D009211:Myoglobin; C010857:apomyoglobin; D009569:Nitric Oxide; D006418:Heme; C029400:heme d1; D009572:Nitrite Reductases; C028123:nitrite reductase (NO-forming)