Total biosynthesis of hydrocortisone from a simple carbon source in yeast.
We report on the production of hydrocortisone, the major adrenal glucocorticoid of mammals and an important intermediate of steroidal drug synthesis, from a simple carbon source by recombinant Saccharomyces cerevisiae strains. An artificial and fully self-sufficient biosynthetic pathway involving 13 engineered genes was assembled and expressed in a single yeast strain. Endogenous sterol biosynthesis was rerouted to produce compatible sterols to serve as substrates for the heterologous part of the pathway. Biosynthesis involves eight mammalian proteins (mature forms of CYP11A1, adrenodoxin (ADX), and adrenodoxin reductase (ADR); mitochondrial forms of ADX and CYP11B1; 3beta-HSD, CYP17A1, and CYP21A1). Optimization involved modulating the two mitochondrial systems and disrupting of unwanted side reactions associated with ATF2, GCY1, and YPR1 gene products. Hydrocortisone was the major steroid produced. This work demonstrates the feasibility of transfering a complex biosynthetic pathway from higher eukaryotes into microorganisms.
Szczebara
Florence Ménard
FM
Laboratoire d'Ingénierie des Protéines Membranaires, CGM du CNRS, F-91198 Gif sur Yvette, France.
Chandelier
Cathy
C
Villeret
Coralie
C
Masurel
Amélie
A
Bourot
Stéphane
S
Duport
Catherine
C
Blanchard
Sophie
S
Groisillier
Agnès
A
Testet
Eric
E
Costaglioli
Patricia
P
Cauet
Gilles
G
Degryse
Eric
E
Balbuena
David
D
Winter
Jacques
J
Achstetter
Tilman
T
Spagnoli
Roberto
R
Pompon
Denis
D
Dumas
Bruno
B
D000818:Animals; D002244:Carbon / Q000378:metabolism*; D002417:Cattle; D003001:Cloning, Molecular; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism; D000431:Ethanol / Q000378:metabolism; D015966:Gene Expression Regulation, Fungal; D005818:Genetic Engineering / Q000379:methods*; D005947:Glucose / Q000378:metabolism; D006801:Humans; D006854:Hydrocortisone / Q000096:biosynthesis* / Q000235:genetics; D009097:Multienzyme Complexes / Q000235:genetics / Q000378:metabolism; D011786:Quality Control; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D011995:Recombination, Genetic; D012441:Saccharomyces cerevisiae / Q000145:classification / Q000235:genetics* / Q000378:metabolism*; D013045:Species Specificity
D009097:Multienzyme Complexes; D011994:Recombinant Proteins; D000431:Ethanol; D002244:Carbon; D003577:Cytochrome P-450 Enzyme System; D005947:Glucose; D006854:Hydrocortisone