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US-20260125731-A1 - MICROORGANISMS AND METHODS FOR PRODUCING CAROTENOIDS AND OTHER COMPOUNDS

US20260125731A1US 20260125731 A1US20260125731 A1US 20260125731A1US-20260125731-A1

Abstract

Recombinant microorganisms configured for enhanced production of carotenoids and other compounds and methods of using the recombinant microorganisms for the production of same. The recombinant microorganism can include a modification that decreases phytoene synthase (CrtB) activity, a modification that decreases lycopene-forming phytoene desaturase (CrtI) activity, a modification that decreases lycopene cyclase (CrtY) activity, a modification that decreases beta-carotene hydroxylase (CrtZ) activity, a modification that decreases 2,2′-beta hydroxylase (CrtG) activity, and/or a modification that increases beta-carotene ketolase (CrtW) activity. The recombinant microorganism can be from the genus Novosphingobium , such as Novosphingobium aromaticivorans.

Inventors

  • Timothy Donohue
  • Daniel Noguera
  • Benjamin Hall
  • Wayne Kontur
  • Derek Gille
  • Jeanette Neri

Assignees

  • WISCONSIN ALUMNI RESEARCH FOUNDATION

Dates

Publication Date
20260507
Application Date
20251028

Claims (20)

  1. 1 . A recombinant microorganism comprising one or more modifications with respect to a corresponding microorganism not comprising the one or more modifications, wherein the one or more modifications comprise any one or more of: a modification that decreases phytoene synthase (CrtB) activity with respect to the corresponding microorganism; a modification that decreases lycopene-forming phytoene desaturase (CrtI) activity with respect to the corresponding microorganism; a modification that decreases lycopene cyclase (CrtY) activity with respect to the corresponding microorganism; a modification that decreases beta-carotene hydroxylase (CrtZ) activity with respect to the corresponding microorganism; a modification that decreases 2,2′-beta hydroxylase (CrtG) activity with respect to the corresponding microorganism; and a modification that increases beta-carotene ketolase (CrtW) activity with respect to the corresponding microorganism.
  2. 2 . The recombinant microorganism of claim 1 , wherein: the modification that decreases phytoene synthase phytoene synthase (CrtB) activity comprises a modification to a gene encoding a phytoene synthase phytoene synthase (CrtB) comprising a sequence having at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% sequence identity to SEQ ID NO:2; the modification that decreases lycopene-forming phytoene (CrtI) desaturase activity comprises a modification to a gene encoding a lycopene-forming phytoene desaturase (CrtI) comprising a sequence having at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% sequence identity to SEQ ID NO:4; the modification that decreases lycopene cyclase (CrtY) activity comprises a modification to a gene encoding a lycopene cyclase (CrtY) comprising a sequence having at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% sequence identity to SEQ ID NO:6; the modification that decreases beta-carotene hydroxylase (CrtZ) activity comprises a modification to a gene encoding a beta-carotene hydroxylase (CrtZ) comprising a sequence having at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% sequence identity to SEQ ID NO:8; and/or the modification that decreases 2,2′-beta hydroxylase (CrtG) activity comprises a modification to a gene encoding a 2,2′-beta hydroxylase (CrtG) comprising a sequence having at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% sequence identity to SEQ ID NO:10.
  3. 3 . The recombinant microorganism of claim 1 , comprising any one or more of: the modification that decreases phytoene synthase phytoene synthase (CrtB) activity; the modification that decreases lycopene-forming phytoene desaturase (CrtI) activity; the modification that decreases lycopene cyclase (CrtY) activity; the modification that decreases beta-carotene hydroxylase (CrtZ) activity; and the modification that decreases 2,2′-beta hydroxylase (CrtG) activity.
  4. 4 . (canceled)
  5. 5 . The recombinant microorganism of claim 1 , comprising the modification that decreases phytoene synthase phytoene synthase (CrtB) activity with respect to the corresponding microorganism.
  6. 6 . The recombinant microorganism of claim 5 , wherein the recombinant microorganism exhibits increased accumulation of coenzyme Q 10 (CoQ 10 ) with respect to the corresponding microorganism.
  7. 7 . The recombinant microorganism of claim 1 , comprising the modification that decreases lycopene cyclase (CrtY) activity.
  8. 8 . The recombinant microorganism of claim 7 , wherein the recombinant microorganism exhibits increased accumulation of lycopene with respect to the corresponding microorganism.
  9. 9 . The recombinant microorganism of claim 1 , comprising the modification that decreases 2,2′-beta hydroxylase (CrtG) activity.
  10. 10 . The recombinant microorganism of claim 9 , wherein the recombinant microorganism exhibits increased accumulation of zeaxanthin with respect to the corresponding microorganism.
  11. 11 . The recombinant microorganism of claim 9 , comprising the modification that decreases beta-carotene hydroxylase (CrtZ) activity.
  12. 12 . The recombinant microorganism of claim 11 , wherein the recombinant microorganism exhibits increased accumulation of beta-carotene with respect to the corresponding microorganism.
  13. 13 . The recombinant microorganism of claim 9 , comprising the modification that increases beta-carotene ketolase (CrtW) activity.
  14. 14 . The recombinant microorganism of claim 13 , wherein the modification that increases beta-carotene ketolase (CrtW) activity comprises a recombinant gene encoding a beta-carotene ketolase (CrtW) comprising a sequence having at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% sequence identity to SEQ ID NO: 12 or SEQ ID NO:14.
  15. 15 . The recombinant microorganism of claim 13 , wherein the recombinant microorganism exhibits increased accumulation of at least one of adonixanthin and astaxanthan with respect to the corresponding microorganism.
  16. 16 . The recombinant microorganism of claim 1 , further comprising: a modification that decreases 2-pyrone-4,6-dicarboxylic acid (PDC) hydrolase (LigI) activity with respect to the corresponding microorganism; a modification that decreases 4-carboxy-2-hydroxy-6-methoxy-6-oxohexa-2,4-dienoate (CHMOD) methyl esterase (DesC) activity with respect to the corresponding microorganism; a modification that decreases 4-carboxy-2-hydroxy-6-methoxy-6-oxohexa-2,4-dienoate (CHMOD) cis-trans isomerase (DesD) activity with respect to the corresponding microorganism; and/or a modification that decreases vanillate/3-O-methylgallate O-demethylase (DmtS) activity with respect to the corresponding microorganism.
  17. 17 . (canceled)
  18. 18 . The recombinant microorganism of claim 16 , wherein the recombinant microorganism exhibits increased accumulation of 2-pyrone-4,6-dicarboxylic acid (PDC) with respect to the corresponding microorganism.
  19. 19 . The recombinant microorganism of claim 1 , wherein the recombinant microorganism is from the genus Novosphingobium.
  20. 20 . The recombinant microorganism of claim 1 , wherein the recombinant microorganism is Novosphingobium aromaticivorans.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS Priority is hereby claimed to U.S. Provisional Application 63/712,785, filed Oct. 28, 2024, which is incorporated herein by reference in its entirety. STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH This invention was made with government support under DE-SC0018409 awarded by the US Department of Energy. The government has certain rights in the invention. SEQUENCE LISTING The instant application contains a Sequence Listing which has been submitted in XML format and is hereby incorporated by reference in its entirety. The XML copy, created on Aug. 22, 2025, is named USPTO-250822-09824639-P230405US02-SEQ_LIST.xml and is 56,670 bytes in size. FIELD OF THE INVENTION The invention is directed to recombinant microorganisms configured for enhanced production of carotenoids and other compounds and methods of using the recombinant microorganisms for the production of same. BACKGROUND Carotenoids are lipophilic isoprenoids that are produced by some plants, algae, bacteria, and fungi, and function as membrane-bound light-harvesting pigments and antioxidants (8, 9). Several carotenoids (including astaxanthin, β-carotene, lycopene, and zeaxanthin) are used industrially as animal feed, food coloring, nutritional supplements, cosmetics additives, and pharmaceuticals, with a 2017 global market size of ˜$1.5B (10, 11). Most industrial carotenoids are produced synthetically (9, 10, 12), though there are a few biological sources commercially being used, such as the flower Tagetes erecta for lutein and the alga Dunaliella salina for 3-carotene (10). Thus, there is growing interest in developing new biological sources of carotenoids (9, 10). New sources for the production of carotenoids such as astaxanthin, β-carotene, lycopene, and zeaxanthin are needed. SUMMARY OF THE INVENTION One aspect of the invention is directed to recombinant microorganisms comprising one or more modifications with respect to a corresponding microorganism not comprising the one or more modifications. In some versions, the one or more modifications comprise any one or more of: a modification that decreases phytoene synthase (CrtB) activity with respect to the corresponding microorganism; a modification that decreases lycopene-forming phytoene desaturase (CrtI) activity with respect to the corresponding microorganism; a modification that decreases lycopene cyclase (CrtY) activity with respect to the corresponding microorganism; a modification that decreases beta-carotene hydroxylase (CrtZ) activity with respect to the corresponding microorganism; a modification that decreases 2,2′-beta hydroxylase (CrtG) activity with respect to the corresponding microorganism; and a modification that increases beta-carotene ketolase (CrtW) activity with respect to the corresponding microorganism. In some versions, the modification that decreases phytoene synthase (CrtB) activity comprises a modification to a gene encoding a phytoene synthase (CrtB) comprising a sequence having at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% sequence identity to SEQ ID NO:2. In some versions, the modification that decreases lycopene-forming phytoene (CrtI) desaturase activity comprises a modification to a gene encoding a lycopene-forming phytoene desaturase (CrtI) comprising a sequence having at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% sequence identity to SEQ ID NO:4. In some versions, the modification that decreases lycopene cyclase (CrtY) activity comprises a modification to a gene encoding a lycopene cyclase (CrtY) comprising a sequence having at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% sequence identity to SEQ ID NO:6. In some versions, the modification that decreases beta-carotene hydroxylase (CrtZ) activity comprises a modification to a gene encoding a beta-carotene hydroxylase (CrtZ) comprising a sequence having at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% sequence identity to SEQ ID NO:8. In some versions, the modification that decreases 2,2′-beta hydroxylase (CrtG) activity comprises a modification to a gene encoding a 2,2′-beta hydroxylase (CrtG) comprising a sequence having at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% sequence identity to SEQ ID NO:10. In some versions, the modification that increases beta-carotene ketolase (CrtW) activity comprises a recombinant gene encoding a beta-carotene ketolase (CrtW) comprising a sequence having at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% sequence identity to SEQ ID NO:12 or SEQ ID NO:14. In some versions, the recombinant microorganism comprises any one or more of: a modification that decreases phytoene synthase (CrtB) activity; a modification that decreases lycopene-forming phytoene desaturase (CrtI) activity; a modification that decreases lycopene cyclase (CrtY) activity; a modification that decreases beta-carotene hydroxylase (CrtZ)