Purification and characterization of NADH oxidase from the archaea Sulfolobus acidocaldarius and Sulfolobus solfataricus

Mariorosario Masullo, Gennaro Raimo, Antonio Dello Russo, Vincenzo Bocchini, Joe V. Bannister

Research output: Contribution to journalArticle

22 Citations (Scopus)

Abstract

The enzyme NADH oxidase (EC 1.6.99.3) has been isolated from the two thermoacidophilic archaea Sulfolobus acidocaldarius and Sulfolobus solfataricus and characterized. In both organisms the enzyme oxidizes specifically β-NADH in the presence of molecular oxygen and requires the presence of a flavin cofactor, showing a high specificity for FAD. A stoicheiometric amount of hydrogen peroxide to NADH is formed as the end product of the reaction, indicating that both enzymes are two-electron donors. The purified enzymes exhibit quite different molecular properties. S. acidocaldarius NADH oxidase is a monomeric protein with an estimated molecular mass of about 27 kDa, whereas S. solfataricus NADH oxidase is a dimeric protein with a molecular mass of 35 kDa per subunit; S. solfataricus NADH oxidase is purified as an FAD-containing protein, whereas S. acidocaldarius NADH oxidase does not contain a flavin molecule. Furthermore, a comparison of the N-terminal amino acid sequence shows no similarities either between the two proteins or to any other NADH oxidases. Both enzymes are essentially thermophilic. In the temperature range 20-80°C, the energy of activation is almost the same for both activities, suggesting that similar energetic parameters are required. Also both oxidases display a great stability to heat. The half-life of heat inactivation is about 180 min at 90°C for S. acidocaldarius NADH oxidase and 77 min at 98°C for the S. solfataricus enzyme. The activity of the two enzymes is inhibited by urea and guanidine and are regulated very differently by several organic solvents.

Original languageEnglish
Pages (from-to)47-54
Number of pages8
JournalBiotechnology and Applied Biochemistry
Volume23
Issue number1
Publication statusPublished - 1996

Fingerprint

Sulfolobus acidocaldarius
Sulfolobus solfataricus
Archaea
Purification
Enzymes
Flavin-Adenine Dinucleotide
Proteins
Molecular mass
NAD
Hot Temperature
Molecular oxygen
Protein S
Guanidine
NADH oxidase
Oxidoreductases
Organic solvents
Hydrogen Peroxide
Half-Life
Urea
Amino Acid Sequence

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Biochemistry
  • Biotechnology

Cite this

Purification and characterization of NADH oxidase from the archaea Sulfolobus acidocaldarius and Sulfolobus solfataricus. / Masullo, Mariorosario; Raimo, Gennaro; Dello Russo, Antonio; Bocchini, Vincenzo; Bannister, Joe V.

In: Biotechnology and Applied Biochemistry, Vol. 23, No. 1, 1996, p. 47-54.

Research output: Contribution to journalArticle

Masullo, Mariorosario ; Raimo, Gennaro ; Dello Russo, Antonio ; Bocchini, Vincenzo ; Bannister, Joe V. / Purification and characterization of NADH oxidase from the archaea Sulfolobus acidocaldarius and Sulfolobus solfataricus. In: Biotechnology and Applied Biochemistry. 1996 ; Vol. 23, No. 1. pp. 47-54.
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