Fe-superoxide dismutase and 2-hydroxy-1,4-benzoquinone reductase preclude the auto-oxidation step in 4-aminophenol metabolism by Burkholderia sp. strain AK-5.
Burkholderia sp. strain AK-5 converts 4-aminophenol to maleylacetic acid via 1,2,4-trihydroxybenzene, which is unstable in vitro and non-enzymatically auto-oxidized to 2-hydroxy-1,4-benzoquinone. Crude extract of strain AK-5 retarded the auto-oxidation and reduced the substrate analogue, 2,6-dimethoxy-1,4-benzoquinone, in the presence of NADH. The two enzymes responsible were purified to homogeneity. The deduced amino acid sequence of the enzyme that inhibited the auto-oxidation showed a high level of identity to sequences of iron-containing superoxide dismutases (Fe-SODs) and contained a conserved metal-ion-binding site; the purified enzyme showed superoxide dismutase activity and contained 1 mol of Fe per mol of enzyme, identifying it as Fe-SOD. Among three type SODs tested, Fe-SOD purified here inhibited the auto-oxidation most efficiently. The other purified enzyme showed a broad substrate specificity toward benzoquinones, including 2-hydroxy-1,4-benzoquinone, converting them to the corresponding 1,4-benzenediols; the enzyme was identified as 2-hydroxy-1,4-benzoquinone reductase. The deduced amino acid sequence did not show a high level of identity to that of benzoquinone reductases from bacteria and fungi that degrade chlorinated phenols or nitrophenols. The indirect role of Fe-SOD in 1,2,4-trihydroxybenzene metabolism is probably to scavenge and detoxify reactive species that promote the auto-oxidation of 1,2,4-trihydroxybenzene in vivo. The direct role of benzoquinone reductase would be to convert the auto-oxidation product back to 1,2,4-trihydroxybenzene. These two enzymes together with 1,2,4-trihydroxybenzene 1,2-dioxygenase convert 1,2,4-trihydroxybenzene to maleylacetic acid.
Takenaka
Shinji
S
Department of Applied Biological Chemistry, Kobe University, Japan. hakko3@kobe-u.ac.jp
Koshiya
Jyun
J
Okugawa
Susumu
S
Takata
Akiko
A
Murakami
Shuichiro
S
Aoki
Kenji
K
D000627:Aminophenols / Q000378:metabolism*; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D001673:Biodegradation, Environmental; D019117:Burkholderia / Q000737:chemistry / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D009153:Mutagens / Q000378:metabolism*; D010084:Oxidation-Reduction; D011808:Quinone Reductases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D013379:Substrate Specificity; D013482:Superoxide Dismutase / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*
D000627:Aminophenols; D001426:Bacterial Proteins; D009153:Mutagens; D013482:Superoxide Dismutase; C100348:1,4-benzoquinone reductase; D011808:Quinone Reductases; C026729:4-aminophenol