Nitrite reductase mechanism

The involvement of HA during bacterial conversion of nitrate to NH3 (known also as the nitrate ammonification phase of the nitrogen cycle) has been studied at the molecular level as part of an effort to delineate the mechanism of conversion of nitrite to NH3 by a group of multiheme cytochromes of bacterial origin. The overall reduction reaction is depicted in equation 3 for cytochrome c-nitrite reductase " ... 

Blom was the first to demonstrate, in 1928, the formation of HA by an unknown mixture of bacteria which utilized nitrate as their sole nitrogen source to produce ammonia , an observation substantiated by Lindsey and Rhines who generalized this reaction to a diverse set of microorganisms capable of producing NH3 by reduction of both nitrites and nitrates. The mechanism of the 6-electron reduction of nitrite to ammonia (i.e. conversion of the [N + 02] species to by bacterial cytochrome c nitrite reductase... 

In spite of the above speculation, the actual mechanisms used by NO-pro-ducing nitrite reductases for reduction of nitrite and its activation for nitrosyl transfer are poorly understood. The fact that both the heme and Cu types of enzyme catalyze the reaction is remarkable in view of the fact that nitrosyl compounds of Fe complexes are well known, whereas Cu-nitrosyl compounds are rare [see Garber and Hollocher (1982) and Kim and Hollocher (1984) for further discussion]. On the other hand, both Fe and Cu can coordinate O and this might be more relevant for the activation of an O atom of nitrite for N-O bond breaking. In the case of the Cu-type nitrite reductase of A. cycloclastes, the... 

Jackson, M. A., Tiedje, J. M., and Averill, B. A. (1991). Evidence for an NO-rebound mechanism for production of N2O from nitrite by the copper-containing nitrite reductase from Achromobacter cyclolastes. FEBS Lett. 291, 41-44. 

Fig. 5. Proposed mechanism for reduction of nitrite by the heme cd-i containing nitrite reductases.

Similar mechanisms operate in the action of nitrate reductase and nitrite reductase. Both of these substances are produced from ammonia by oxidation. Plants and soil bacteria can reduce these compounds to provide ammonia for metabolism. The common agricultural fertilizer ammonium nitrate, NH4NO3, provides reduced nitrogen for plant growth directly, and by providing a substrate for nitrate reduction. NADH or NADPH is the electron donor for nitrate reductase, depending on the organism. 

This chapter focuses on the chemistry ofbiomimetic copper nitrosyl complexes relevant to the NO-copper interactions in proteins that are central players in dissimilatory nitrogen oxide reduction (denitrification). The current state of knowledge of NO-copper interactions in nitrite reductase, a key denitrifying enzyme, is briefly surveyed the syntheses, structures, and reactivity of copper nitrosyl model complexes prepared to date are presented and the insight these model studies provide into the mechanisms of denitrification and the structures of other copper protein nitrosyl intermediates are discussed. Emphasis is placed on analysis of the geometric features, electronic structures, and biomimetic reactivity with NO or NOf of the only structurally characterized copper nitrosyls, a dicopper(II) complex bridged by NO and a mononuclear tris(pyrazolyl)hydroborate complex having a Cu(I)-NO formulation. 

Cutruzzola, F., Arese, M., Grasso, S., Bellelli, A., and Brunori, M., 1997, Tyrosine 10 in the c-haem domain is not involved in the catalytic mechanism of nitrite reductase from Pseudomonas aeruginosa, FEBS Lett. 412 3659369. 

Figure 6 Proposed mechanisms for copper-containing nitrite reductase (a) via an O-bonded nitrosyP or (b) a bound HONO intermediate ...

Cytochrome cd nitrite reductase from Paracoceus pantotrophus has a different mechanism, with two identical subunits, each with domains containing a c-type cytochrome heme and a dj-type cytochrome heme. Electrons from external donors enter through the c heme the d heme is the site of nitrite reduction to NO and oxygen reduction to water. One of the puzzles of the mechanism is how the NO can escape from... 

FIGURE 16-14 Proposed Mechanism of Nitrite Reductase. (Redrawn from M. J. Boulanger,... 

Still another nitrite reductase, cytochrome c NIR, contains five heme groups, only one of which functions as the active site. A combination of calculations and crystallographic studies has suggested a mechanism in which nitrite replaces a water molecule on one side of the Fe(II) heme (a lysine N is on the opposite side), one of the oxygens of N02 is protonated, and the N — 0 bond is broken with loss of H2O, leaving a linear Fe(III)—NO species with a low-spin Fe(III). Addition of two electrons and H" " leads to Fe HNO, which is then reduced to Fe H2NOH. Yet another electron and another allow release of H2O and formation of an Fe NH3 complex. Release of ammonia and a final electron and water addition complete the cycle. Overall, six electrons and seven hydrogen ions react with the nitrite ... 

Direct reaction of NO with enzymes has been shown for cytochrome c oxidase (cyt c oxidase). The reaction of NO with the binuclear metal centre of cyt c oxidase apparently leads to the formation of nitrite at the active site [123] the mechanism of which was described as the opposite of nitrite reduction to NO by non-haem nitrite reductases [124]. The inhibition was caused by the binding of NO to the reduced copper centre of the enzyme rather than the expected reaction with Fe. ... 

Mechanism. The electron-transport pathway of nitrite reductase begins with the type 1 copper center of pseudoazurin, continues to the type 1 copper center of nitrite reductase, and from there to its type 2 copper center. This last electron transfer is not conducted directly from copper to copper, but via an intramolecular Cysl36-Hisl35 bridge, similar to the one proposed for ascorbate oxidase [26,208] (Fig. 35). 

Figure 41 Proposed mechanism of nitrite reductase. Reproduced from E. T. Adman M. E. P. Murphy, in Handbook of Metalloproteins A. Messerschmidt, R. Huber, T. Pouios, K. Wieghardt, Eds. John Wiley Sons Chichester, 2001 Vol. 2, pp 1381-1390, with permission from John Wiley Sons.

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