The one electron reduction of nitrite to NO, catalyzed in vivo by a group of enzymes called nitrite reductases (NIRs), has been extensively investigated due to its relevance in the processes of denitrification. The heme containing NIRs are Soluble noncovalent homodimers with two heme groups in each subunit: a d(1) heme, the site of nitrite reduction, and a type c heme, responsible for internal electron transfer. High resolution X-ray structures show that all NIRs have two distal histidines in the active site as a common feature. We analyzed the reaction mechanism of nitrite reduction by Pseudomonas aeruginosa (Pa) NIR using a combined quantum mechanics/molecular mechanics (QM-MM) approach. The central Fe(II) porphyrin complex plus the proximal and distal histidines were treated at the density functional theory level, while the solvated protein environment was modeled using the Amber force field. Our results indicate that nitrite binds to the reduced active site with one histidine protonated and that, after protonation of the second histidine, proton transfer and dehydration result in an Fe-III(NO) species plus a free water molecule coordinated to both histidines. The computed results also suggest that the NO ligand is probably displaced by another water molecule in the distal cavity. The enzyme finally recovers the resting state, after proton reorganization, reduction, and water formation. Our results show that the catalytic capacity of the enzyme lies primarily on the distal histidines, real guards of the active site cavity.