Genetic engineering of Bacillus megaterium for high-yield production of the major teleost progestogens 17α,20ß-di- and 17α,20ß,21α-trihydroxy-4-pregnen-3-one.
Metab Eng
; 36: 19-27, 2016 07.
Article
en En
| MEDLINE
| ID: mdl-26976492
ABSTRACT
17α,20ß-Dihydroxy-4-pregnen-3-one (17α,20ßDiOH-P) and 17α,20ß,21α-trihydroxy-4-pregnen-3-one (20ßOH-RSS) are the critical hormones required for oocyte maturation in fish. We utilized B. megaterium's endogenous 20ß-hydroxysteroid dehydrogenase (20ßHSD) for the efficient production of both progestogens after genetically modifying the microorganism to reduce side-product formation. First, the gene encoding the autologous cytochrome P450 CYP106A1 was deleted, resulting in a strain devoid of any steroid hydroxylation activity. Cultivation of this strain in the presence of 17α-hydroxyprogesterone (17αOH-P) led to the formation of 17α,20α-dihydroxy-4-pregnen-3-one (17α,20αDiOH-P) as a major and 17α,20ßDiOH-P as a minor product. Four enzymes were identified as 20αHSDs and their genes deleted to yield a strain with no 20αHSD activity. The 3-oxoacyl-(acyl-carrier-protein) reductase FabG was found to exhibit 20ßHSD-activity and overexpressed to create a biocatalyst yielding 0.22g/L 17α,20ßDiOH-P and 0.34g/L 20ßOH-RSS after 8h using shake-flask cultivation, thus obtaining products that are at least a thousand times more expensive than their substrates.
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MEDLINE
Asunto principal:
Progestinas
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Bacillus megaterium
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Mejoramiento Genético
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Redes y Vías Metabólicas
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Vías Biosintéticas
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Ingeniería Metabólica
Límite:
Animals
Idioma:
En
Año:
2016
Tipo del documento:
Article