WO2014100251A1 - Voie de biodésulfuration basée sur le cytochrome p450 - Google Patents

Voie de biodésulfuration basée sur le cytochrome p450 Download PDF

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Publication number
WO2014100251A1
WO2014100251A1 PCT/US2013/076254 US2013076254W WO2014100251A1 WO 2014100251 A1 WO2014100251 A1 WO 2014100251A1 US 2013076254 W US2013076254 W US 2013076254W WO 2014100251 A1 WO2014100251 A1 WO 2014100251A1
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WO
WIPO (PCT)
Prior art keywords
engineered
microbial
pathway
variant
cell
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Ceased
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PCT/US2013/076254
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English (en)
Inventor
Pedro D. COELHO
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California Institute of Technology
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California Institute of Technology
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Publication date
Application filed by California Institute of Technology filed Critical California Institute of Technology
Publication of WO2014100251A1 publication Critical patent/WO2014100251A1/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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Classifications

    • C—CHEMISTRY; METALLURGY
    • C10—PETROLEUM, GAS OR COKE INDUSTRIES; TECHNICAL GASES CONTAINING CARBON MONOXIDE; FUELS; LUBRICANTS; PEAT
    • C10G—CRACKING HYDROCARBON OILS; PRODUCTION OF LIQUID HYDROCARBON MIXTURES, e.g. BY DESTRUCTIVE HYDROGENATION, OLIGOMERISATION, POLYMERISATION; RECOVERY OF HYDROCARBON OILS FROM OIL-SHALE, OIL-SAND, OR GASES; REFINING MIXTURES MAINLY CONSISTING OF HYDROCARBONS; REFORMING OF NAPHTHA; MINERAL WAXES
    • C10G27/00—Refining of hydrocarbon oils in the absence of hydrogen, by oxidation
    • C—CHEMISTRY; METALLURGY
    • C10—PETROLEUM, GAS OR COKE INDUSTRIES; TECHNICAL GASES CONTAINING CARBON MONOXIDE; FUELS; LUBRICANTS; PEAT
    • C10G—CRACKING HYDROCARBON OILS; PRODUCTION OF LIQUID HYDROCARBON MIXTURES, e.g. BY DESTRUCTIVE HYDROGENATION, OLIGOMERISATION, POLYMERISATION; RECOVERY OF HYDROCARBON OILS FROM OIL-SHALE, OIL-SAND, OR GASES; REFINING MIXTURES MAINLY CONSISTING OF HYDROCARBONS; REFORMING OF NAPHTHA; MINERAL WAXES
    • C10G32/00—Refining of hydrocarbon oils by electric or magnetic means, by irradiation, or by using microorganisms
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/0004—Oxidoreductases (1.)
    • C12N9/0012—Oxidoreductases (1.) acting on nitrogen containing compounds as donors (1.4, 1.5, 1.6, 1.7)
    • C12N9/0036—Oxidoreductases (1.) acting on nitrogen containing compounds as donors (1.4, 1.5, 1.6, 1.7) acting on NADH or NADPH (1.6)
    • C12N9/0038—Oxidoreductases (1.) acting on nitrogen containing compounds as donors (1.4, 1.5, 1.6, 1.7) acting on NADH or NADPH (1.6) with a heme protein as acceptor (1.6.2)
    • C12N9/0042—NADPH-cytochrome P450 reductase (1.6.2.4)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/0004—Oxidoreductases (1.)
    • C12N9/0071—Oxidoreductases (1.) acting on paired donors with incorporation of molecular oxygen (1.14)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Y—ENZYMES
    • C12Y106/00—Oxidoreductases acting on NADH or NADPH (1.6)
    • C12Y106/02—Oxidoreductases acting on NADH or NADPH (1.6) with a heme protein as acceptor (1.6.2)
    • C12Y106/02004—NADPH-hemoprotein reductase (1.6.2.4), i.e. NADP-cytochrome P450-reductase
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Y—ENZYMES
    • C12Y114/00—Oxidoreductases acting on paired donors, with incorporation or reduction of molecular oxygen (1.14)
    • C12Y114/14—Oxidoreductases acting on paired donors, with incorporation or reduction of molecular oxygen (1.14) with reduced flavin or flavoprotein as one donor, and incorporation of one atom of oxygen (1.14.14)
    • C12Y114/14001—Unspecific monooxygenase (1.14.14.1)

Definitions

  • DszA, DszB, DszC, DszD Four enzymes (DszA, DszB, DszC, DszD) are involved in catalyzing the four oxidative reactions in the 4S pathway ( Figure 2), which is found in diverse microbes including species of Rhodococcus, Bacillus, Gordonia, Paenibacillus, Pseudomonas and Mycobacterium, among others.
  • the first enzyme, dibenzothiophene monooxygenase (DszC) catalyzes two sequential oxidations converting DBT to dibenzothiophene sulfoxide (DBTO) and dibenzothiophene sulfone (DBT0 2 ).
  • T268A hinders compound I formation
  • T268A, and other mutations at position 268 for example, substitutions that do not contain a side chain having a hydroxyl, including substitutions of G, L, I, V, or other non-polar or hydrophobic amino acid
  • mutation of amino acids having a side chain in its 3-dimensional vicinity e.g., 5 or 10 Angstrom radius
  • Such mutations may be included in mutation libraries and screened in E. coli as described above.
  • the engineered P450 replaces DscZ, the first step in the pathway. See Figure 2.
  • the microbe expresses a biodesulfurization metabolic pathway: the metabolic pathway comprising P450 or variant thereof (including a variant described herein), a dibenzothiophene sulfone monooxygenase, 2-hydroxybiphenyl-2-sulfmate sulfino lyase, and a flavin reductase.
  • the metabolic pathway may convert dibenzothiophene (DBT) compounds to 2- hydroxybiphenyl-2-sulfinate (HBPS), HBPS to 2-hydroxybiphenyl (HBP), and HBP to sulfite.
  • DBT dibenzothiophene
  • HBPS 2- hydroxybiphenyl-2-sulfinate
  • HBP 2-hydroxybiphenyl
  • HBP 2-hydroxybiphenyl

Landscapes

  • Chemical & Material Sciences (AREA)
  • Organic Chemistry (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Zoology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Wood Science & Technology (AREA)
  • Genetics & Genomics (AREA)
  • General Health & Medical Sciences (AREA)
  • Biochemistry (AREA)
  • General Engineering & Computer Science (AREA)
  • Oil, Petroleum & Natural Gas (AREA)
  • Microbiology (AREA)
  • Molecular Biology (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • General Chemical & Material Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • Biomedical Technology (AREA)
  • Biotechnology (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)

Abstract

L'invention concerne des catalyseurs à base de cytochrome P450 modifiés pour catalyser l'oxydation de soufre aromatique en sulfoxydes et sulfones. L'invention concerne en outre des catalyseurs microbiens modifiés pour retirer du pétrole le thiophène, le benzothiophène, le dibenzothiophène et leurs dérivés, et des procédés de biodésulfuration du pétrole.
PCT/US2013/076254 2012-12-18 2013-12-18 Voie de biodésulfuration basée sur le cytochrome p450 Ceased WO2014100251A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US201261738863P 2012-12-18 2012-12-18
US61/738,863 2012-12-18

Publications (1)

Publication Number Publication Date
WO2014100251A1 true WO2014100251A1 (fr) 2014-06-26

Family

ID=50979169

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US2013/076254 Ceased WO2014100251A1 (fr) 2012-12-18 2013-12-18 Voie de biodésulfuration basée sur le cytochrome p450

Country Status (1)

Country Link
WO (1) WO2014100251A1 (fr)

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN110241095A (zh) * 2019-06-05 2019-09-17 西南医科大学 一种cyp119酶及其突变体和用途

Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6100074A (en) * 1994-11-03 2000-08-08 Bg Plc Mutant mono-oxygenase cytochrome P-450 cam
US20090209010A1 (en) * 2006-08-04 2009-08-20 California Institute Of Technology Methods and systems for selective fluorination of organic molecules
US20110086405A1 (en) * 2007-09-27 2011-04-14 Taijiro Tomikawa Method for producing hydroxylated adamantane using cytochrome p450
US7960155B1 (en) * 1999-07-27 2011-06-14 Basf Aktiengesellschaft Cytochrome p450 monooxygenases and their use for oxidizing organic compounds

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6100074A (en) * 1994-11-03 2000-08-08 Bg Plc Mutant mono-oxygenase cytochrome P-450 cam
US7960155B1 (en) * 1999-07-27 2011-06-14 Basf Aktiengesellschaft Cytochrome p450 monooxygenases and their use for oxidizing organic compounds
US20090209010A1 (en) * 2006-08-04 2009-08-20 California Institute Of Technology Methods and systems for selective fluorination of organic molecules
US20110086405A1 (en) * 2007-09-27 2011-04-14 Taijiro Tomikawa Method for producing hydroxylated adamantane using cytochrome p450

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
KIRIMURA, KOHTARO ET AL.: "Identification and functional analysis of the genes encoding dibenzothiophene-desulfurizing enzymes from thermophilic bacteria", APPLIED MICROBIOLOGY AND BIOTECHNOLOGY, vol. 65, no. 6, November 2004 (2004-11-01), pages 703 - 713 *

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN110241095A (zh) * 2019-06-05 2019-09-17 西南医科大学 一种cyp119酶及其突变体和用途
CN110241095B (zh) * 2019-06-05 2022-11-01 西南医科大学 一种cyp119酶及其突变体和用途

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