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ES2621657T3 - Polinucleótidos que codifican enzimas modificadoras de isoprenoides y procedimientos de uso de los mismos - Google Patents

Polinucleótidos que codifican enzimas modificadoras de isoprenoides y procedimientos de uso de los mismos Download PDF

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ES2621657T3
ES2621657T3 ES12158465.0T ES12158465T ES2621657T3 ES 2621657 T3 ES2621657 T3 ES 2621657T3 ES 12158465 T ES12158465 T ES 12158465T ES 2621657 T3 ES2621657 T3 ES 2621657T3
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nucleotide sequence
seq
set forth
nucleic acid
amino acid
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Dae-Kyun Ro
Karyn Newman
Eric M. Paradise
Jay D. Keasling
Mario Ouellet
Rachel Eachus
Kimberly Ho
Timothy Ham
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University of California Berkeley
University of California San Diego UCSD
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University of California San Diego UCSD
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    • 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
    • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
    • C12P17/00—Preparation of heterocyclic carbon compounds with only O, N, S, Se or Te as ring hetero atoms
    • C12P17/18—Preparation of heterocyclic carbon compounds with only O, N, S, Se or Te as ring hetero atoms containing at least two hetero rings condensed among themselves or condensed with a common carbocyclic ring system, e.g. rifamycin
    • C12P17/181—Heterocyclic compounds containing oxygen atoms as the only ring heteroatoms in the condensed system, e.g. Salinomycin, Septamycin
    • 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
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
    • C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
    • C12N15/82—Vectors or expression systems specially adapted for eukaryotic hosts for plant cells, e.g. plant artificial chromosomes (PACs)
    • C12N15/8241—Phenotypically and genetically modified plants via recombinant DNA technology
    • C12N15/8242—Phenotypically and genetically modified plants via recombinant DNA technology with non-agronomic quality (output) traits, e.g. for industrial processing; Value added, non-agronomic traits
    • C12N15/8243—Phenotypically and genetically modified plants via recombinant DNA technology with non-agronomic quality (output) traits, e.g. for industrial processing; Value added, non-agronomic traits involving biosynthetic or metabolic pathways, i.e. metabolic engineering, e.g. nicotine, caffeine
    • 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

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  • Biophysics (AREA)
  • Plant Pathology (AREA)
  • General Chemical & Material Sciences (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)
  • Preparation Of Compounds By Using Micro-Organisms (AREA)
  • Enzymes And Modification Thereof (AREA)
  • Breeding Of Plants And Reproduction By Means Of Culturing (AREA)

Abstract

Polinucleótido aislado que comprende una secuencia nucleotídica que codifica una citocromo P450 reductasa (CPR), en el que la secuencia nucleotídica codifica un polipéptido que tiene al menos un 90% de identidad de secuencia aminoacídica con la secuencia aminoacídica expuesta en SEQ ID NO: 4, en el que la CPR transfiere electrones desde NADPH a una amorfa-4,11-dieno oxidasa que tiene la secuencia aminoacídica expuesta en SEQ ID NO: 2.

Description

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de la modificación enzimática se analizan generalmente por cromatografía de gases y espectrometría de masas. Si un ácido nucleico objeto codifica una sesquiterpeno-oxidasa o una sesquiterpeno-hidroxilasa puede determinarse fácilmente mediante ensayos estándar para estas actividades enzimáticas. Véase, por ejemplo, la publicación de patente de los EE. UU. n° 20050019882.
5 [0051] En el presente documento se describe un ácido nucleico objeto que comprende la secuencia nucleotídica mostrada en la figura 1 y expuesta en SEQ ID NO: 1. Un ácido nucleico objeto puede comprender una secuencia nucleotídica con al menos aproximadamente el 45%, al menos aproximadamente el 50%, al menos aproximadamente el 55%, al menos aproximadamente el 57%, al menos aproximadamente el 60%, al menos aproximadamente el 65%, al menos aproximadamente el 70%, al menos aproximadamente el 75%, al menos aproximadamente el 80%, al menos aproximadamente el 85%, al menos aproximadamente el 90%, al menos aproximadamente el 95%, al menos aproximadamente el 98% o al menos aproximadamente el 99% de identidad de secuencia nucleotídica con la secuencia nucleotídica expuesta en SEQ ID NO: 1. Un ácido nucleico objeto puede comprender una secuencia nucleotídica con una, dos, tres, cuatro, cinco, seis, siete, ocho, nueve, diez, de
15 aproximadamente 10 a aproximadamente 15, de aproximadamente 15 a aproximadamente 20, de aproximadamente 20 a aproximadamente 25 o de aproximadamente 25 a aproximadamente 50 sustituciones de nucleótidos en comparación con la secuencia nucleotídica expuesta en la SEQ ID NO: 1.
[0052] Un ácido nucleico objeto puede comprender una secuencia nucleotídica con al menos aproximadamente el 45%, al menos aproximadamente el 50%, al menos aproximadamente el 55%, al menos aproximadamente el 57%, al menos aproximadamente el 60%, al menos aproximadamente el 65%, al menos aproximadamente el 70%, al menos aproximadamente el 75%, al menos aproximadamente el 80%, al menos aproximadamente el 85%, al menos aproximadamente el 90%, al menos aproximadamente el 95%, al menos aproximadamente el 98% o al menos aproximadamente el 99% de identidad de secuencia nucleotídica con la
25 secuencia nucleotídica expuesta en SEQ ID NO: 1, en que el ácido nucleico codifica un polipéptido que muestra actividad de terpeno-hidroxilasa y/o terpeno-oxidasa (por ejemplo, actividad de sesquiterpeno-oxidasa, actividad de sesquiterpeno-hidroxilasa, etc.).
[0053] Un ácido nucleico objeto puede comprender una secuencia nucleotídica con al menos aproximadamente el 50%, al menos aproximadamente el 55%, al menos aproximadamente el 57%, al menos aproximadamente el 60%, al menos aproximadamente el 65%, al menos aproximadamente el 70%, al menos aproximadamente el 75%, al menos aproximadamente el 80%, al menos aproximadamente el 85%, al menos aproximadamente el 90%, al menos aproximadamente el 95%, al menos aproximadamente el 98% o al menos aproximadamente el 99% de identidad de secuencia nucleotídica con un tramo de al menos aproximadamente 500,
35 al menos aproximadamente 600, al menos aproximadamente 700, al menos aproximadamente 800, al menos aproximadamente 900, al menos aproximadamente 1.000, al menos aproximadamente 1.100, al menos aproximadamente 1.200, al menos aproximadamente 1.300, al menos aproximadamente 1.400 o al menos aproximadamente 1.450 nucleótidos contiguos de la secuencia nucleotídica expuesta en SEQ ID NO: 1.
[0054] Un ácido nucleico objeto puede comprender al menos aproximadamente 500, al menos aproximadamente 600, al menos aproximadamente 700, al menos aproximadamente 800, al menos aproximadamente 900, al menos aproximadamente 1.000, al menos aproximadamente 1.100, al menos aproximadamente 1.200, al menos aproximadamente 1.300, al menos aproximadamente 1.400 o al menos aproximadamente 1.450 nucleótidos contiguos de la secuencia nucleotídica expuesta en SEQ ID NO: 1. Un ácido
45 nucleico objeto puede comprender al menos aproximadamente 500, al menos aproximadamente 600, al menos aproximadamente 700, al menos aproximadamente 800, al menos aproximadamente 900, al menos aproximadamente 1.000, al menos aproximadamente 1.100, al menos aproximadamente 1.200, al menos aproximadamente 1.300, al menos aproximadamente 1.400 o al menos aproximadamente 1.450 nucleótidos contiguos de la secuencia nucleotídica expuesta en SEQ ID NO: 1 y codifica un polipéptido que muestra actividad de terpeno-hidroxilasa y/o terpeno-oxidasa, por ejemplo, actividad de sesquiterpeno-hidroxilasa y/u oxidasa.
[0055] Un ácido nucleico objeto puede comprender una secuencia nucleotídica que hibrida en condiciones de hibridación restrictivas con un ácido nucleico que comprende la secuencia nucleotídica expuesta en SEQ ID NO: 1 o un complemento de ésta.
55 [0056] En el presente documento se describe un ácido nucleico objeto que comprende una secuencia nucleotídica que codifica un polipéptido que comprende una secuencia aminoacídica según se muestra en la figura 2 y se expone en SEQ ID NO: 2. Un ácido nucleico objeto puede comprender una secuencia nucleotídica que codifica un polipéptido que comprende una secuencia aminoacídica con al menos aproximadamente el 45%, al menos aproximadamente el 50%, al menos aproximadamente el 55%, al menos aproximadamente el 60%, al menos aproximadamente el 65%, al menos aproximadamente el 70%, al menos aproximadamente el 75%, al menos aproximadamente el 80%, al menos aproximadamente el 85%, al menos aproximadamente el 90%, al menos aproximadamente el 95%, al menos aproximadamente el 98% o al menos aproximadamente el 99% de identidad de secuencia aminoacídica con la secuencia aminoacídica expuesta en SEQ ID NO: 2. Un ácido nucleico objeto puede
65 comprender una secuencia nucleotídica que codifica un polipéptido que comprende una secuencia aminoacídica con al menos aproximadamente el 45%, al menos aproximadamente el 50%, al menos aproximadamente el 55%, al
10
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Claims (1)

  1. imagen1
ES12158465.0T 2005-07-05 2006-06-29 Polinucleótidos que codifican enzimas modificadoras de isoprenoides y procedimientos de uso de los mismos Active ES2621657T3 (es)

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US697067P 2005-07-05

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EP (2) EP1919514B1 (es)
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PT1919514E (pt) 2012-08-16
EP2489672B1 (en) 2017-01-18
HK1134495A1 (en) 2010-04-30
CN101495499B (zh) 2013-09-04
CN101495499A (zh) 2009-07-29
CN103667370B (zh) 2016-08-10
AP2013006962A0 (en) 2013-07-31
BRPI0612411B1 (pt) 2020-05-05
WO2007005604A3 (en) 2008-12-31
PL1919514T3 (pl) 2012-10-31
US20100218283A1 (en) 2010-08-26
EP1919514A4 (en) 2009-12-02
KR101444090B1 (ko) 2014-09-26
US20120288905A1 (en) 2012-11-15
CA2613469A1 (en) 2007-01-11
DK1919514T3 (da) 2012-08-20
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CN106939323B (zh) 2021-07-13
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