Reductive dehalogenation by cytochrome P450CAM: Substrate binding and catalysis

Journal Article · Tue Sep 14 00:00:00 EDT 1993 · Biochemistry; (United States)

DOI:https://doi.org/10.1021/bi00087a014· OSTI ID:5941580

Li, S; Wackett, L P ^[1]

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1. Univ. of Minnesota, St. Paul (United States)

Biological reductive dehalogenation reactions are important in environmental detoxification of organohalides. Only scarce information is available on the enzymology underlying these reactions. Cytochrome P450CAM with a known X-ray structure and well-studied oxygenase reaction cycle, has been studied for its ability to reduce carbon-halogen bonds under anaerobic conditions. The reductive reactions functioned with NADH and the physiological electron-transfer proteins or by using artificial electron donors to reduce cytochrome P450CAM. Halogenated methane and ethane substrates were transformed by a two-electron reduction and subsequent protonation, beta-elimination, or alpha-elimination to yield alkanes, alkene, or carbene-derived products, respectively. Halogenated substrates bound to the camphor binding site as indicated by saturable changes in the Fe(III)-heme spin state upon substrate addition. Hexachloromethane was bound with a dissociation constant (KD) of 0.7 microM and caused > 95% shift from low- to high-spin iron. Ethanes bearing fewer chlorine substituents were bound with increasing dissociation constants and gave lesser degrees of iron spin-state change. Camphor competitively inhibited hexachloroethane reduction with an inhibitor constant (KI) similar to the dissociation constant for camphor (KI = KD = 0.9 microM). Rate determinations with pentachloroethane indicated a 100-fold higher enzyme V/K compared to the second-order rate constant for hematin free in solution. These studies on substrate binding and catalysis will help reveal how biological systems enzymatically reduce carbon-halogen bonds in the environment.

• OSTI ID: 5941580
• Journal Information: Biochemistry; (United States), Vol. 32:36; ISSN 0006-2960
• Country of Publication: United States
• Language: English

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
CYTOCHROMES
BIOCHEMICAL REACTION KINETICS
HALOGENATED ALIPHATIC HYDROCARBONS
DEHALOGENATION
PSEUDOMONAS
ENZYME ACTIVITY
CARBON DIOXIDE
CATALYSIS
DETOXIFICATION
ELECTRON TRANSFER
FREONS
HEME
HYDROXYLASES
NAD
BACTERIA
CARBON COMPOUNDS
CARBON OXIDES
CARBOXYLIC ACIDS
CHALCOGENIDES
CHEMICAL REACTIONS
COENZYMES
ENZYMES
HETEROCYCLIC ACIDS
HETEROCYCLIC COMPOUNDS
KINETICS
MICROORGANISMS
NUCLEOTIDES
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC HALOGEN COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
OXIDES
OXIDOREDUCTASES
OXYGEN COMPOUNDS
PIGMENTS
PORPHYRINS
PROTEINS
REACTION KINETICS
550200* - Biochemistry
550700 - Microbiology