Abstract
Nitrite oxidoreductase, the essential enzyme complex of nitrite oxidizing membranes, was isolated from cells of the nitrifying bacterium Nitrobacter hamburgensis. The enzyme system was solubilized and purified in the presence of 0.25% sodium deoxycholate. Nitrite oxidoreductase oxidized nitrite to nitrate in the presence of ferricyanide. The pH optimum was 8.0, and the apparent K m value for nitrite amounted to 3.6 mM. With reduced methyl-and benzylviologen nitrite oxidoreductase exhibited nitrate reductase activity with an apparent K m value of 0.9 mM for nitrate. NADH was also a suitable electron donor for nitrate reduction. The pH optimum was 7.0.
Treatment with SDS resulted in the dissociation into 3 subunits of 116,000, 65,000 and 32,000. The enzyme complex contained iron, molydbenum, sulfur and copper. A c-type cytochrome was present. Isolated nitrite oxidoreductase is a particle of 95±30 Å in diameter.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
Abbreviations
- DOC:
-
sodium deoxycholate
References
Aleem MIH (1968) Mechanism of oxidative phosphorylation in the chemoautotroph Nitrobacter agilis. Biochim Biophys Acta 162:338–447
Aleem MIH, Sewell DL (1981) Mechanism of nitrite oxidation and oxidoreductase systems in Nitrobacter agilis. Curr Microbiol 5:267–272
Aleem MIH, Hoch GE, Varner JE (1965) Water as the source of oxidant and reductant in bacterial chemosynthesis. Proc Natl Acad Sci USA 54:869–873
Alef K, Klemme JH (1979) Assimilatory nitrate reductase of Rhodopseudomonas capsulata AD2: a molybdo hemoprotein. Z Naturforsch 34c:33–37
Barton GM (1965) α, α′-Dipyridyl as a phenol detecting reagent. J Chromato 20:189
Beuscher N, Mayer F, Gottschalk G (1974) Citrate lyase from Rhodospeudomonas gelatinosa: purification, electron microscopy and subunit structure. Arch Microbiol 100:308–328
Bligh EG, Dyer WJ (1959) A rapid method of total lipid extraction and purification. Can J Biochem Physiol 37:911–917
Bock E, Heinrich G (1969) Morphologische und physiologische Untersuchungen an Zellen von Nitrobacter winogradskyi Buch. Arch Mikrobiol 69:149–159
Bock E, Sundermeyer-Klinger H, Stackebrandt E (1983) New facultative lithoautotrophic nitrite-oxidizing bacteria. Arch Microbiol 136:281–284
Cobley GJ (1976a) Energy-conserving reactions in phosphorylating electron transport particles from Nitrobacter winogradskyi. Biochem J 156:481–491
Cobley GJ (1976b) Reduction of cytochromes by nitrite in electron particles from Nitrobacter winogradskyi. Biochem J 156:493–498
Estabrook RW (1961) Studies of oxidative phosphorylation with potassium ferricyanide as electron acceptor. J Biol Chem 236:3051–3056
Gazotti P, Bock E, Fleischer S (1975) Interaction of D-β-hydroxybutyrate apodehydrogenase with phospholipids. J Biol Chem 250:5782–5790
Heubült J (1929) Untersuchungen über Nitritbakterien. Planta 8:398–422
Hewitt EJ, Notton BA (1980) Nitrate reductase systems in eukaryotic and prokaryotic organisms. In: Coughlan MP (ed) Molybdenum and molybdenum containing enzymes Pergamon Press, Oxford, pp 275–325
Kiesow L (1964) On the assimilation of energy source from inorganic sources in autotrophic forms of life. Proc Natl Acad Sci USA 52:980–988
Kumar S, Nicholas DJD, Williams EH (1983) Definite 15N NMR evidence that water serves as source of ‘0’ during nitrite oxidation by Nitrobacter agilis. FEBS Lett 152:71–74
Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T 4. Nature (Lond) 227:685–690
Layne E (1957) Spectrometric and turbidometric methods for measuring proteins. In: Colowick SP, Kaplan NO (eds) Methods in enzymology, vol III. Academic Press, New York, pp 447–554
Lees H, Simpson JR (1957) The biochemistry of the nitrifying organisms. 5. Nitrite oxidation by Nitrobacter. Biochem J 65:297–305
Marshall Stewart JC (1980) Colorimetric determination of phospholipids with ammonium ferrothiocyanate. Anal Biochem 104:10–14
Milde K, Bock E (1984) Isolation and partial characterization of inner and outer membrane fractions of Nitrobacter hamburgensis. FEMS Microbiol Lett 21:137–141
Remsen CC, Watson SW (1972) Freeze-etching of bacteria. Intern Rev Cytol, vol 33. Academic Press, New York London, pp 253–296
Satoh T (1981) Soluble dissimilatory nitrate reductase containing cytochrome c from a photodenitrifier, Rhodopseudomonas shaeroides forma sp. denitrificans. Plant Cell Physiol 22:443–452
Schramel P, Klose B-J (1981) Direktbestimmung von Cu, Fe, Zn, Ca, Mg, und Na im Serum mittels ICP-Emissionsspektralanalyse. Fresenius Z Anal Chem 307:26–30
Schramel P, Klose B-J (1982) ICP-Atomemissionsspektroskopie, eine neue Analysentechnik zum Nachweis von Spurenelementen in biologischen Materialien. In: Welz B (ed) Atomspektrometrische Spurenanalytik. Verlag Chemie, Weinheim, pp 75–79
Singer SJ, Nicholson GL (1973) The fluid mosaic model of the structure of cell membranes. Science 175:720–731
Steinmüller W, Bock E (1976) Growth of Nitrobacter in the presence of organic matter. I. Mixotrophic growth. Arch Microbiol 108:299–304
Stouthamer AH (1976) Biochemistry and genetics of nitrate reductase in bacteria. In: Rose AH, Tempest DW (eds) Adv Microbiol Physiol 14. Academic Press, New York London, pp 315–375
Sundermeyer H, Bock E (1981) Characterization of the nitriteoxidizing system in Nitrobacter. In: Bothe H, Trebst A (eds) Biology of inorganic nitrogen and sulfur. Springer, Berlin Heidelberg New York, pp 317–324
Tanaka Y, Fukumori Y, Yamanaka T (1983) Purification of cytochrome a1c1 from Nitrobacter agilis and characterization of nitrite oxidation system of the bacterium. Arch Microbiol 135:265–271
Tanford C, Reynolds JA (1976) Characterization of membrane proteins in detergent solutions. Biochim Biophys Acta 457:133–170
Tsien HC, Lambert R, Laudelout H (1968) Fine structure and the localization of the nitrite oxidizing system in Nitrobacter winogradski. Antonie van Leeuwenhoek. J Microbiol 34:483–494
Valentine RC, Shapiro BM, Stadtman ET (1968) Regulation of glutamine synthetase. XII. Electron microscopy of the enzyme from Escherichia coli. Biochemistry 7:2143
Veslemøy E, Anderson M, Odd van Frøholm L, Laane MM (1980) Isolation of a relatively pure outer membrane fraction of Moraxella nonliquiefaciens and a comparison of its characteristics with the cytoplasmic membrane-containing material. J Gen Microbiol 116:149–155
Winogradsky S (1890) Recherches sur les organismes de la nitrification. Ann Inst Pasteur 4:213–231
Yamanaka T, Kamita Y, Fukumori Y (1982) Molecular and enzymatic properties of “cytochrome aa3”-type terminal oxidase derived from Nitrobacter agilis. J Biochem 89:265–273
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Sundermeyer-Klinger, H., Meyer, W., Warninghoff, B. et al. Membrane-bound nitrite oxidoreductase of Nitrobacter: evidence for a nitrate reductase system. Arch. Microbiol. 140, 153–158 (1984). https://doi.org/10.1007/BF00454918
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00454918