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 language | English |
|---|---|
| Pages (from-to) | 47-54 |
| Number of pages | 8 |
| Journal | Biotechnology and Applied Biochemistry |
| Volume | 23 |
| Issue number | 1 |
| Publication status | Published - 1996 |
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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 journal › Article
}
TY - JOUR
T1 - Purification and characterization of NADH oxidase from the archaea Sulfolobus acidocaldarius and Sulfolobus solfataricus
AU - Masullo, Mariorosario
AU - Raimo, Gennaro
AU - Dello Russo, Antonio
AU - Bocchini, Vincenzo
AU - Bannister, Joe V.
PY - 1996
Y1 - 1996
N2 - 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.
AB - 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.
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UR - http://www.scopus.com/inward/citedby.url?scp=0030063350&partnerID=8YFLogxK
M3 - Article
C2 - 8867896
AN - SCOPUS:0030063350
VL - 23
SP - 47
EP - 54
JO - Biotechnology and Applied Biochemistry
JF - Biotechnology and Applied Biochemistry
SN - 0885-4513
IS - 1
ER -