Nova Patents
CA2602183C

Production of carotenoids in oleaginous yeast and fungi

Abstract

The present invention provides systems for producing engineered oleaginous yeast or fungi that express carotenoids.

CA2602183C, drawing sheet 1
Sheet 1 of 31

Term

No projected expiry on record.

  1. Priority
  2. Filed
  3. Granted
  4. Today

20 claims: 10 independent, 10 dependent

  1. 1
    CA 02602183 2013-10-29 What is claimed is:1. A recombinant Yarrowia fungus, wherein said recombinant fungus: (a) is oleaginous in that it can accumulate lipid to at least 20% of its dry cell weight;and (b) produces at least one carotenoid, and can accumulate the produced carotenoid to at least 1% of its dry cell weight, wherein said fungus comprises at least one modification which is a carotenogenic modification, an oleaginic modification, or both, wherein said carotenogenic modification comprises a genetic modification resulting in increased production of one or more carotenoids, wherein said oleaginic modification comprises a genetic modification resulting in at least one of increased production and activity of one or more oleaginic polypeptides, and wherein said at least one modification alters oleaginicity of said recombinant fungus, confers to said recombinant fungus oleaginy, confers to said recombinant fungus the ability to produce the at least one carotenoid to a level at least 1% of its dry cell weight, or confers to said recombinant fungus the ability to produce at least one carotenoid which the fungus does not naturally produce.
  2. 4
    The recombinant fungus of any one of claims 1 to 3, wherein the at least one oleaginic modification comprises expression of at least one heterologous gene encoding the at least one heterologous oleaginic polypeptide. CA 02602183 2013-10-29
  3. 6
    The recombinant fungus of any one of claims 1 to 5, wherein the at least one carotenogenic modification increases or decreases expression of at least one carotenogenic polypeptide which is:(i) an isoprenoid biosynthesis polypeptide which is: acetoacetyl-CoA thiolase, HMG-CoA synthase, HMG-CoA reductase, mevalonate kinase, phosphomevalonate kinase, mevalonate pyrophosphate decarboxylase, IPP isomerase, FPP synthase, or GGPP synthase;(ii) a carotenoid biosynthesis polypeptide which is: a phytoene synthase, phytoene dehydrogenase, lycopene cyclase, carotenoid ketolase, carotenoid hydroxylase, astaxanthin synthase, carotenoid epsilon hydroxylase, carotenoid glucosyltransferase, lycopene cyclase (beta and epsilon subunits), or acyl CoA:diacyglycerol acyltransferase;(iii) an isoprenoid biosynthesis competitor polypeptides which is: squalene synthase, prenyldiphosphate synthase, or PHB polyprenyltransferase;(iv) a carotenogenic polypeptide which is any one of the polypeptides of Tables 7-25, 29, and 30, or any combination thereof;or (v) any combination of (i) to (iv).
  4. 10
    The recombinant fungus of any one of claims 1 to 9, wherein the same accumulates the produced at least one carotenoid to a level which is:above 1%, above 2%, above 3%, above 5%, or above 10% of the fungus1 dry cell weight.
  5. 11
    The recombinant fungus of any one claims 1 to 10, characterized in that the same accumulates lipid in the form of cytoplasmic bodies.
  6. 13
    The recombinant fungus of any one of claims 1 to 12, wherein said fungus comprises a carotenogenic modification which is:(a) expression of a polypeptide which is: a truncated endogenous HMG CoA reductase polypeptide that lacks the N-terminal membrane spanning domain, acetoacetyl-CoA thiolase, HMG-CoA synthase, FPP synthase, or GGPP synthase;(b) expression of a heterologous polypeptide which is: phytoene synthase, phytoene desaturase, lycopene cyclase, carotenoid ketolase, carotenoid hydroxylase, or any combination thereof;(c) decreased expression or activity of an endogenous polypeptide which is: squalene synthase, prenyldiphosphate synthase, PHB polyprenyltransferase;or (d) any combination of (a) to (c).
  7. 15
    A method of producing a carotenoid, the method comprising:(a) cultivating the recombinant fungus as defined in any one of claims 1 to 14 under conditions that allow production of the carotenoid;and (b) isolating said produced carotenoid.
  8. 18
    The method of any one of claims 15 to 17, wherein the carotenoid is astaxanthin, β-carotene, canthaxanthin, zeaxanthin, lutein, lycopene, or any combination thereof.
  9. 19
    The method of any one of claims 15 to 18, wherein the carotenoid comprises astaxanthin. 101 CA 02602183 2013-10-29
  10. 20
    A method of preparing a food or feed additive containing a carotenoid, the method comprising:(a) cultivating the recombinant fungus as defined in any one of claims 1 to 14 under conditions that allow production of the carotenoid;(b) isolating the carotenoid;and (c) combining the isolated carotenoid with one or more other food or feed additive components. 102
Independent claims10