Abstract
A cytoplasmic NADH oxidase (NOX) was purified from a soil bacteria, Brevibacterium sp. KU1309, which is able to grow in the medium containing 2-phenylethanol as the sole source of carbon under an aerobic condition. The enzyme catalyzed the oxidation of NADH to NAD+ involving two-electron reduction of O2 to H2O2. The molecular weight of the enzyme was estimated to be 102 kDa by gel filtration and 57 kDa by SDS-PAGE, which indicates that the NOX was a homodimer consisting of a single subunit. The enzyme was stable up to 70°C at a broad range of pH from 7 to 11. The enzyme activity increased about ten-fold with the addition of ammonium salt, while it was inhibited by Zn2+ (39%), Cu2+ (41%), Hg2+ (72%) and Ag+ (37%). The enzyme acts on NADH, but not on NADPH. The regeneration of NAD+ utilizing this enzyme made selective oxidation of mandelic acid or l-phenylalanine possible. This thermostable enzyme is expected to be applicable as a useful biocatalyst for NAD+ recycling.
| Original language | English |
|---|---|
| Pages (from-to) | 71-78 |
| Number of pages | 8 |
| Journal | Applied Microbiology and Biotechnology |
| Volume | 80 |
| Issue number | 1 |
| DOIs | |
| Publication status | Published - 2008 Aug |
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Keywords
- Brevibacterium sp.
- Cofactor regeneration
- Hydrogen peroxide
- Molecular oxygen
- NADH oxidase
ASJC Scopus subject areas
- Biotechnology
- Microbiology
- Bioengineering
- Microbiology (medical)
Cite this
Purification and characterization of thermostable H2O 2-forming NADH oxidase from 2-phenylethanol-assimilating Brevibacterium sp. KU1309. / Hirano, Jun Ichiro; Miyamoto, Kenji; Ohta, Hiromichi.
In: Applied Microbiology and Biotechnology, Vol. 80, No. 1, 08.2008, p. 71-78.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Purification and characterization of thermostable H2O 2-forming NADH oxidase from 2-phenylethanol-assimilating Brevibacterium sp. KU1309
AU - Hirano, Jun Ichiro
AU - Miyamoto, Kenji
AU - Ohta, Hiromichi
PY - 2008/8
Y1 - 2008/8
N2 - A cytoplasmic NADH oxidase (NOX) was purified from a soil bacteria, Brevibacterium sp. KU1309, which is able to grow in the medium containing 2-phenylethanol as the sole source of carbon under an aerobic condition. The enzyme catalyzed the oxidation of NADH to NAD+ involving two-electron reduction of O2 to H2O2. The molecular weight of the enzyme was estimated to be 102 kDa by gel filtration and 57 kDa by SDS-PAGE, which indicates that the NOX was a homodimer consisting of a single subunit. The enzyme was stable up to 70°C at a broad range of pH from 7 to 11. The enzyme activity increased about ten-fold with the addition of ammonium salt, while it was inhibited by Zn2+ (39%), Cu2+ (41%), Hg2+ (72%) and Ag+ (37%). The enzyme acts on NADH, but not on NADPH. The regeneration of NAD+ utilizing this enzyme made selective oxidation of mandelic acid or l-phenylalanine possible. This thermostable enzyme is expected to be applicable as a useful biocatalyst for NAD+ recycling.
AB - A cytoplasmic NADH oxidase (NOX) was purified from a soil bacteria, Brevibacterium sp. KU1309, which is able to grow in the medium containing 2-phenylethanol as the sole source of carbon under an aerobic condition. The enzyme catalyzed the oxidation of NADH to NAD+ involving two-electron reduction of O2 to H2O2. The molecular weight of the enzyme was estimated to be 102 kDa by gel filtration and 57 kDa by SDS-PAGE, which indicates that the NOX was a homodimer consisting of a single subunit. The enzyme was stable up to 70°C at a broad range of pH from 7 to 11. The enzyme activity increased about ten-fold with the addition of ammonium salt, while it was inhibited by Zn2+ (39%), Cu2+ (41%), Hg2+ (72%) and Ag+ (37%). The enzyme acts on NADH, but not on NADPH. The regeneration of NAD+ utilizing this enzyme made selective oxidation of mandelic acid or l-phenylalanine possible. This thermostable enzyme is expected to be applicable as a useful biocatalyst for NAD+ recycling.
KW - Brevibacterium sp.
KW - Cofactor regeneration
KW - Hydrogen peroxide
KW - Molecular oxygen
KW - NADH oxidase
UR - http://www.scopus.com/inward/record.url?scp=47149101254&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=47149101254&partnerID=8YFLogxK
U2 - 10.1007/s00253-008-1535-x
DO - 10.1007/s00253-008-1535-x
M3 - Article
C2 - 18521590
AN - SCOPUS:47149101254
VL - 80
SP - 71
EP - 78
JO - Applied Microbiology and Biotechnology
JF - Applied Microbiology and Biotechnology
SN - 0175-7598
IS - 1
ER -