PMID- 16820465
OWN - NLM
STAT- MEDLINE
DCOM- 20060905
LR  - 20181113
IS  - 0099-2240 (Print)
IS  - 1098-5336 (Electronic)
IS  - 0099-2240 (Linking)
VI  - 72
IP  - 7
DP  - 2006 Jul
TI  - Purification and characterization of an arene cis-dihydrodiol dehydrogenase 
      endowed with broad substrate specificity toward polycyclic aromatic hydrocarbon 
      dihydrodiols.
PG  - 4726-34
AB  - Initial reactions involved in the bacterial degradation of polycyclic aromatic 
      hydrocarbons (PAHs) include a ring-dihydroxylation catalyzed by a dioxygenase and 
      a subsequent oxidation of the dihydrodiol products by a dehydrogenase. In this 
      study, the dihydrodiol dehydrogenase from the PAH-degrading Sphingomonas strain 
      CHY-1 has been characterized. The bphB gene encoding PAH dihydrodiol 
      dehydrogenase (PDDH) was cloned and overexpressed as a His-tagged protein. The 
      recombinant protein was purified as a homotetramer with an apparent Mr of 
      110,000. PDDH oxidized the cis-dihydrodiols derived from biphenyl and eight 
      polycyclic hydrocarbons, including chrysene, benz[a]anthracene, and 
      benzo[a]pyrene, to corresponding catechols. Remarkably, the enzyme oxidized 
      pyrene 4,5-dihydrodiol, whereas pyrene is not metabolized by strain CHY-1. The 
      PAH catechols produced by PDDH rapidly auto-oxidized in air but were regenerated 
      upon reaction of the o-quinones formed with NADH. Kinetic analyses performed 
      under anoxic conditions revealed that the enzyme efficiently utilized two- to 
      four-ring dihydrodiols, with Km values in the range of 1.4 to 7.1 microM, and 
      exhibited a much higher Michaelis constant for NAD+ (Km of 160 microM). At pH 
      7.0, the specificity constant ranged from (1.3 +/- 0.1) x 10(6) M(-1) s(-1) with 
      benz[a]anthracene 1,2-dihydrodiol to (20.0 +/- 0.8) x 10(6) M(-1) s(-1) with 
      naphthalene 1,2-dihydrodiol. The catalytic activity of the enzyme was 13-fold 
      higher at pH 9.5. PDDH was subjected to inhibition by NADH and by 
      3,4-dihydroxyphenanthrene, and the inhibition patterns suggested that the 
      mechanism of the reaction was ordered Bi Bi. The regulation of PDDH activity 
      appears as a means to prevent the accumulation of PAH catechols in bacterial 
      cells.
FAU - Jouanneau, Yves
AU  - Jouanneau Y
AD  - CEA, DSV, Departement de Reponse et Dynamique Cellulaires/BBSI, CEA-Grenoble, 17 
      avenue des Martyrs, F-38054 Grenoble Cedex 9, France. yves.jouanneau@cea.fr
FAU - Meyer, Christine
AU  - Meyer C
LA  - eng
SI  - GENBANK/AM230449
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PL  - United States
TA  - Appl Environ Microbiol
JT  - Applied and environmental microbiology
JID - 7605801
RN  - 0 (Naphthalenes)
RN  - 0 (Polycyclic Aromatic Hydrocarbons)
RN  - 0 (trans-1,2-dihydro-1,2-naphthalenediol)
RN  - 2166IN72UN (naphthalene)
RN  - EC 1.- (Oxidoreductases)
SB  - IM
MH  - Cloning, Molecular
MH  - Escherichia coli/enzymology/genetics
MH  - Kinetics
MH  - Molecular Sequence Data
MH  - Naphthalenes/chemistry/*metabolism
MH  - *Oxidoreductases/chemistry/genetics/isolation & purification/metabolism
MH  - Polycyclic Aromatic Hydrocarbons/chemistry/*metabolism
MH  - Sequence Analysis, DNA
MH  - Sphingomonas/*enzymology/genetics
MH  - Substrate Specificity
PMC - PMC1489356
EDAT- 2006/07/06 09:00
MHDA- 2006/09/06 09:00
PMCR- 2006/11/01
CRDT- 2006/07/06 09:00
PHST- 2006/07/06 09:00 [pubmed]
PHST- 2006/09/06 09:00 [medline]
PHST- 2006/07/06 09:00 [entrez]
PHST- 2006/11/01 00:00 [pmc-release]
AID - 72/7/4726 [pii]
AID - 0395-06 [pii]
AID - 10.1128/AEM.00395-06 [doi]
PST - ppublish
SO  - Appl Environ Microbiol. 2006 Jul;72(7):4726-34. doi: 10.1128/AEM.00395-06.