Mycobacterium smegmatis is a suitable cell factory for the production of steroidic synthons.
A number of pharmaceutical steroid synthons are currently produced through the microbial side-chain cleavage of natural sterols as an alternative to multi-step chemical synthesis. Industrially, these synthons have been usually produced through fermentative processes using environmental isolated microorganisms or their conventional mutants. Mycobacterium smegmatis mc2155 is a model organism for tuberculosis studies which uses cholesterol as the sole carbon and energy source for growth, as other mycobacterial strains. Nevertheless, this property has not been exploited for the industrial production of steroidic synthons. Taking advantage of our knowledge on the cholesterol degradation pathway of M. smegmatis mc2155 we have demonstrated that the MSMEG_6039 (kshB1) and MSMEG_5941 (kstD1) genes encoding a reductase component of the 3-ketosteroid 9α-hydroxylase (KshAB) and a ketosteroid Δ1-dehydrogenase (KstD), respectively, are indispensable enzymes for the central metabolism of cholesterol. Therefore, we have constructed a MSMEG_6039 (kshB1) gene deletion mutant of M. smegmatis MS6039 that transforms efficiently natural sterols (e.g. cholesterol and phytosterols) into 1,4-androstadiene-3,17-dione. In addition, we have demonstrated that a double deletion mutant M. smegmatis MS6039-5941 [ΔMSMEG_6039 (ΔkshB1) and ΔMSMEG_5941 (ΔkstD1)] transforms natural sterols into 4-androstene-3,17-dione with high yields. These findings suggest that the catabolism of cholesterol in M. smegmatis mc2155 is easy to handle and equally efficient for sterol transformation than other industrial strains, paving the way for valuating this strain as a suitable industrial cell factory to develop à la carte metabolic engineering strategies for the industrial production of pharmaceutical steroids.
Department of Environmental Biology, Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas, Ramiro de Maeztu 9, 28040, Madrid, Spain.
Beatriz
B
Galán
Department of Environmental Biology, Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas, Ramiro de Maeztu 9, 28040, Madrid, Spain.|MRC Centre for Molecular Bacteriology and Infection, Department of Medicine, Imperial College London, London, SW7 2AZ, UK.
Iria
I
Uhía
Department of Environmental Biology, Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas, Ramiro de Maeztu 9, 28040, Madrid, Spain.
Esther
E
García-Fernández
Department of Environmental Biology, Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas, Ramiro de Maeztu 9, 28040, Madrid, Spain.
Igor
I
Martínez
Department of Biotechnology, Gadea Biopharma, Parque Tecnológico de León, Nicostrato Vela s/n, 24009, León, Spain.
Esther
E
Bahíllo
Department of Biotechnology, Gadea Biopharma, Parque Tecnológico de León, Nicostrato Vela s/n, 24009, León, Spain.
Juan L
JL
de la Fuente
Department of Biotechnology, Gadea Biopharma, Parque Tecnológico de León, Nicostrato Vela s/n, 24009, León, Spain.
José L
JL
Barredo
Department of Environmental Biology, Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas, Ramiro de Maeztu 9, 28040, Madrid, Spain.
Lorena
L
Fernández-Cabezón
Department of Environmental Biology, Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas, Ramiro de Maeztu 9, 28040, Madrid, Spain.
José L
JL
García
D001711:Biotransformation; D017353:Gene Deletion; D005798:Genes, Bacterial; D053858:Metabolic Networks and Pathways / Q000235:genetics*; D020102:Mycobacterium smegmatis / Q000235:genetics* / Q000378:metabolism*; D013261:Sterols / Q000378:metabolism*
D013261:Sterols