Achromobacter nitrite reductase

The blue protein from A. faecalis strain S-6, which was isolated as a requirement for transferring electrons to a copper-containing nitrite reductase, has since been shown to have sequence homology with proteins arbitrarily designated pseudoazurin by Ambler and Tobari (1985), from Achromobacter cycloclastes and from Pseudomonas AMI. [Pseudomonas AMI also produces amicyanin, which is the recipient of electrons from methylamine dehydrogenase, (see below)]. In A. cycloclastes reduced pseudoazurin donates electrons to a copper nitrite reductase (Liu et ai, 1986), as it does in A. faecalis. Ambler and Tobari (1985)... 

The nature of the copper in these proteins is not totally clear. Dooley et al (1988) reported that the Achromobacter protein may have two kinds of type I sites in a total of three copper sites per dimeric protein, while the A. faecalis protein was reported to be a tetrameric protein with both type I and type II coppers (KakutanielaZ., 1981). Interestingly, the Achromobacter protein is green. Both of these nitrite reductases accept electrons from a cupredoxin. 

Fenderson, F. F., Kumar, S., Adman, E. T., Liu, M.-Y., Payne, W. J., and LeGall,]. (1991). Amino acid sequence of nitrite reductase A copper protein from Achromobacter cyclo-clastes. Biochemistry 30, 7180-7185. 

Hulse, C. L., and Averill, B. A. (1989). Evidence for a copper-nitrosyl intermediate in denitrification by the copper-containing nitrite reductase of Achromobacter cycloclastes. J. Am. Chem. Soc. Ill, 2322-2323. 

Iwasaki, N., Noji, S., and Shidara, S. (1975). Achromobacter cycloclastes nitrite reductase. The function of copper, amino acid composition, and ESR spectra. J. Biochem. (Tokyo) 78, 355-361. 

Libby, E., and Averill, B. A. (1992). Evidence that the type 2 copper centers are the site of nitrite reduction by Achromobacter cycloclastes nitrite reductase. Biochem. Biophys. Res. Commun. 187, 1529-1535. 

A denitrifying nitrite reductase from Achromobacter cycloclastes1533 and Ps. denitrificans1534 is a copper protein, containing two moles copper per mole of protein (molecular weight 69 000). This reduces nitrite to nitrogen monoxide. 

The crystal structure of nitrite reductase (NiR) from Achromobacter cycloclastes was recently reported by Godden et al. (46). The protein contains both a type I and a type II copper site. The type I center has a distorted tetrahedral structure typical of type I copper, whereas the... 

Figure 1. Schematic representation of the copper sites in nitrite reductase from Achromobacter cycloclastes (17).

The Physical Properties of Purified Nitrite Reductase FROM Achromobacter fiseheri"... 

The nitrite reductase system of Achromobacter fischeri appears to be composed of two separable enzymes (341). The first enzyme is a flavin reductase and utilizes NADH or NADPH to reduce FMN or FAD. The second interacts with the flavin reductase and converts nitrite and hy-droxylaraine to ammonia. The nitrite reductase enzyme has a molecular weight of 95,000 4,000 (Table XVII), contains two heme c per mole, and is inhibited by p-mercuribenzoate, cyanide, and carbon monoxide. 

VII According to Husain and Sadana (433), the earlier preparation of Achromobacter fischeri nitrite reductase with a molecular weight of... 

Achromobacter fischeri, nitrite reductase, physical properties, 277-279 Active site, lipoamide dehydrogenase, 105 Acyl hydrazides, catalase and, 379 Acyltransferase activity, glyceraldehyde-... 

Figure 3 Electronic absorption (A) and magnetic circular dichroism (B) spectra of the Type 1 Cu site of Achromobacter cycloclastes nitrite reductase (a) and spinach plastocyanin (b). Absorption data were obtained at 120 K for nitrite reductase and at 25 K for plastocyanin. MCD spectra were obtained at 4.2 K. Gaussian resolution of bands in the absorption spectra is based on a simultaneous linear least-squares fit of Abs, MCD, and CD data for each. MCD data from 5000 to 8000 cm have been multiplied by a factor of 5. (Reprinted with permission from Ref. 22. 1996 American Chemical Society)...

Fig. 8. (a) Drawing of the trimer of nitrite reductase from Achromobacter cycloclastes. (b) Drawing of the interface between domain 1 (subunit A) and domain 2 of the adjacent symmetry-related molecule (subunit C) of nitrite reductase from A. cycloclastes. (c) Drawing of domain 1 and 3 of ascorbate oxidase. The type-1 copper is in domain 3 and the trinuclear copper center is between domain 1 and domain 3. The domains have an orientation similar to that of the corresponding domains of the nitrite reductase shown in b. The figure was produced by the RIBBON Program (S7). 

Welinder, A.C., Zhang, J., Hansen, A.G., Moth-Poulsen, K., Christensen, H.E.M., Kuznetsov, A.M., Bjornholm, T., and Ulstrup, J. (2007) Voltammetry and electrocatalysis of achromobacter xylosoxidans copper nitrite reductase on functionalized Au(lll) electrode surfaces. Zeitschriji jur Physikalische Chemie-International Journal of Research in Physical Chemistry ej Chemical Physics, 221,1343-1378. 

Fia. 39. The absorption spectra of Achromobacter fischeri nitrite reductase. Spectra were recorded in 0.05 M phosphate buffer, pH 7.5, at 0.41 mg enzyme protcin/ml. 

VII According to Husain and Sadana (4S3), the earlier preparation of Achromobacter fischeri nitrite reductase with a molecular weight of 95,000 40,000 341) was found to be polydisperse. A monodisperse preparation subsequently studied (433) had a molecular weight of 80,000, and was shown to be composed of two subunits of approximate molecular weights of 39,000 with methionine as the sole N-terminal residue. The subunits are stated to be linked together by disulfide bridges. 

Robust voltammetry and in situ STM to molecular resolution have been achieved when the Au(lll)-electrode surfaces are modified by linker molecules, Fig. 8-10, prior to protein adsorption. Comprehensive voltammetric data are available for horse heart cyt and P. aeruginosa The latter protein, which we address in the next Section, has in a sense emerged as a paradigm for nanoscale bioelectrochemistry. We address first briefly two other proteins, viz. the electron transfer iron-sulfur protein Pyrococcus furiosus ferredoxin and the redox metalloenz5mie Achromobacter xylosoxidans copper nitrite reductase. 

Iwasaki H, Matsubara T (1972) A nitrite reductase from Achromobacter cycloclastes. J Biochem 71 645-652... 

Until now, pseudoazurin has only been found in bacteria, e.g., in the denitrifying bacteria Alcaligenes faecalis and Achromobacter cycloclastes. It is the electron donor of the green copper-protein nitrite reductase which catalyses the reduction of nitrite (NO2) to nitrogen monoxide (NO) [86-89]. The physiological electron donor of pseudoazurin is as yet unknown [70]. Pseudoazurin has a molecular mass of approximately 13.5 kD and a chain length of about 123 amino acid residues [88]. The additional amino acids, as compared with plastocyanin, form the C-terminal end of the protein (Fig. 14). The pseudoazurins have redox potentials of about 230 mV [90,91]. 

Most proteins with type 1 copper centers are blue, although the nitrite reductases from Achromobacter cycloclastes, Alcaligenes faecalis, and Pseudomonas aureofaciens, are green [26,27]. This is probably caused by a distortion of the type 1 copper center, although the interrelation of distortion and absorption properties of the copper centers have not yet been clarified [27]. 

Fig. 34. The surrounding structure of the type 1 and type 2 copper-binding centers of nitrite reductase from Alcaligenes faecalis. The structure is equivalent to that of Achromobacter. From Averill 1996 [26] with permission...

All of the nitrite reductases just described convert NO2" to NO in a dissimilatory manner for energy extraction other nitrite reductases in assimilatory pathways convert N02 directly to NH4+ for synthetic purposes. Most of these are nonheme flavoproteins, but one has been found in Achromobacter fischeri (422) with properties similar to the dissimilatory reductases two hemes c per molecule of weight 95,000. The analogy... 

Nitrite Reductase Achromobacter cycloclastes 458nm = 2200 Esssmn 1.800 1.22 rhombic (73) 2.56 278,279... 

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