Laboratory evolution of a soluble, self-sufficient, highly active alkane hydroxylase

Anton Glieder, Edgardo T Farinas, Frances H Arnold

Research output: Contribution to journalArticleResearchpeer-review

Abstract

We have converted cytochrome P450 BM-3 from Bacillus megaterium (P450 BM-3), a medium-chain (C12-C18) fatty acid monooxygenase, into a highly efficient catalyst for the conversion of alkanes to alcohols. The evolved P450 BM-3 exhibits higher turnover rates than any reported biocatalyst for the selective oxidation of hydrocarbons of small to medium chain length (C3-C8). Unlike naturally occurring alkane hydroxylases, the best known of which are the large complexes of methane monooxygenase (MMO) and membrane-associated non-heme iron alkane monooxygenase (AlkB), the evolved enzyme is monomeric, soluble, and requires no additional proteins for catalysis. The evolved alkane hydroxylase was found to be even more active on fatty acids than wild-type BM-3, which was already one of the most efficient fatty acid monooxgenases known. A broad range of substrates including the gaseous alkane propane induces the low to high spin shift that activates the enzyme. This catalyst for alkane hydroxylation at room temperature opens new opportunities for clean, selective hydrocarbon activation for chemical synthesis and bioremediation.

Original languageEnglish
Pages (from-to)1135-9
Number of pages5
JournalMolecular biotechnology
Volume20
Issue number11
DOIs
Publication statusPublished - Nov 2002

Fingerprint

Cytochrome P-450 CYP4A
Alkanes
Paraffins
Fatty acids
Fatty Acids
methane monooxygenase
Hydrocarbons
Mixed Function Oxygenases
Enzymes
Bacillus megaterium
Propane
Hydroxylation
Catalysts
Environmental Biodegradation
Biocatalysts
Bioremediation
Bacilli
Catalysis
Chain length
Cytochrome P-450 Enzyme System

Keywords

  • Alcohols
  • Alkanes
  • Bacillus megaterium
  • Bacterial Proteins
  • Catalysis
  • Cytochrome P-450 CYP4A
  • Cytochrome P-450 Enzyme System
  • Enzyme Activation
  • Evolution, Molecular
  • Hydroxylation
  • Mixed Function Oxygenases
  • Mutagenesis, Site-Directed
  • Oxidation-Reduction
  • Proteomics
  • Quality Control
  • Recombinant Proteins
  • Sensitivity and Specificity
  • Solubility
  • Species Specificity
  • Journal Article
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

Cite this

Laboratory evolution of a soluble, self-sufficient, highly active alkane hydroxylase. / Glieder, Anton; Farinas, Edgardo T; Arnold, Frances H.

In: Molecular biotechnology, Vol. 20, No. 11, 11.2002, p. 1135-9.

Research output: Contribution to journalArticleResearchpeer-review

Glieder, Anton ; Farinas, Edgardo T ; Arnold, Frances H. / Laboratory evolution of a soluble, self-sufficient, highly active alkane hydroxylase. In: Molecular biotechnology. 2002 ; Vol. 20, No. 11. pp. 1135-9.
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AB - We have converted cytochrome P450 BM-3 from Bacillus megaterium (P450 BM-3), a medium-chain (C12-C18) fatty acid monooxygenase, into a highly efficient catalyst for the conversion of alkanes to alcohols. The evolved P450 BM-3 exhibits higher turnover rates than any reported biocatalyst for the selective oxidation of hydrocarbons of small to medium chain length (C3-C8). Unlike naturally occurring alkane hydroxylases, the best known of which are the large complexes of methane monooxygenase (MMO) and membrane-associated non-heme iron alkane monooxygenase (AlkB), the evolved enzyme is monomeric, soluble, and requires no additional proteins for catalysis. The evolved alkane hydroxylase was found to be even more active on fatty acids than wild-type BM-3, which was already one of the most efficient fatty acid monooxgenases known. A broad range of substrates including the gaseous alkane propane induces the low to high spin shift that activates the enzyme. This catalyst for alkane hydroxylation at room temperature opens new opportunities for clean, selective hydrocarbon activation for chemical synthesis and bioremediation.

KW - Alcohols

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KW - Species Specificity

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