Coexpression of VHb and MceG genes in Mycobacterium sp. Strain LZ2 enhances androstenone production via immobilized repeated batch fermentation

Zhang, Yang; Zhou, Xiuling; Yao, Yingying; Xu, Qianqian; Hai-ying, Shi; Wang, Kuiming; Feng, Wei; Shen, Yanbing

doi:10.1016/j.biortech.2021.125965

Cited by 14 publications

(11 citation statements)

References 45 publications

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“…qRT‐PCR analysis was performed as previously described (Liu et al., 2018 ) on a StepOne™ RT‐qPCR System (Life Technologies, USA) using SYBR Green Master Mix (Yeasen, China). The 16S rRNA expression was used as the internal standard for normalization with 2 −ΔΔCt algorithm (Zhang et al., 2021 ). Three technical replicates were performed to obtain the mean value.…”

Section: Methodsmentioning

confidence: 99%

Microbial metabolism of diosgenin by a novel isolated Mycolicibacterium sp. HK‐90: A promising biosynthetic platform to produce 19‐carbon and 21‐carbon steroids

Wang,

Qiu,

Chen

et al. 2024

Microbial Biotechnology

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Green manufacture of steroid precursors from diosgenin by microbial replacing multistep chemical synthesis has been elusive. It is currently limited by the lack of strain and degradation mechanisms. Here, we demonstrated the feasibility of this process using a novel strain Mycolicibacterium sp. HK‐90 with efficiency in diosgenin degradation. Diosgenin degradation by strain HK‐90 involves the selective removal of 5,6‐spiroketal structure, followed by the oxygenolytic cleavage of steroid nuclei. Bioinformatic analyses revealed the presence of two complete steroid catabolic gene clusters, SCG‐1 and SCG‐2, in the genome of strain HK‐90. SCG‐1 cluster was found to be involved in classic phytosterols or cholesterol catabolic pathway through the deletion of key kstD1 gene, which promoted the mutant m‐∆kstD1 converting phytosterols to intermediate 9α‐hydroxyandrostenedione (9‐OHAD). Most impressively, global transcriptomics and characterization of key genes suggested SCG‐2 as a potential gene cluster encoding diosgenin degradation. The gene inactivation of kstD2 in SCG‐2 resulted in the conversion of diosgenin to 9‐OHAD and 9,16‐dihydroxy‐pregn‐4‐ene‐3,20‐dione (9,16‐(OH)2‐PG) in mutant m‐ΔkstD2. Moreover, the engineered strain mHust‐ΔkstD1,2,3 with a triple deletion of kstDs was constructed, which can stably accumulate 9‐OHAD by metabolizing phytosterols, and accumulate 9‐OHAD and 9,16‐(OH)2‐PG from diosgenin. Diosgenin catabolism in strain mHust‐ΔkstD1,2,3 was revealed as a progression through diosgenone, 9,16‐(OH)2‐PG, and 9‐OHAD to 9α‐hydroxytestosterone (9‐OHTS). So far, this work is the first report on genetically engineered strain metabolizing diosgenin to produce 21‐carbon and 19‐carbon steroids. This study presents a promising biosynthetic platform for the green production of steroid precursors, and provide insights into the complex biochemical mechanism of diosgenin catabolism.

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Section: Methodsmentioning

confidence: 99%

Microbial metabolism of diosgenin by a novel isolated Mycolicibacterium sp. HK‐90: A promising biosynthetic platform to produce 19‐carbon and 21‐carbon steroids

Wang,

Qiu,

Chen

et al. 2024

Microbial Biotechnology

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“…It was concluded that the Mce4 transporter plays an active role in the uptake and transport of sterols. Zhang et al (2021) [ 77 ] coexpressed optimized Vitreoscilla hemoglobin (VHb) and MceG genes in M . sp.…”

Section: Rational Construction Of Recombinant M Neoaurum ...mentioning

confidence: 99%

“…In addition, the overexpression of vgb in Msp-vgb/mceG resulted in an intracellular ROS level of only 69% of that of Msp and had a smaller upward trend. Msp-vgb/mceG has a higher catalase activity than Msp, which may be the main reason why vgb is able to reduce ROS levels and improve strain activity [ 77 ].…”

Section: Rational Construction Of Recombinant M Neoaurum ...mentioning

confidence: 99%

“…Cell immobilization technology has been widely used in the production of biological products as an effective means of overcoming the limitations of mass transfer, improving the stability of biocatalyst operation, protecting cells from toxic substrates or products, improving production efficiency, and reducing production costs [ 115 ]. Zhang et al (2021) [ 77 ] immobilized Msp-vgb/mceG with bagasse as immobilized material. Under the same substrate level, immobilized cell fermentation resulted in higher biomass, yield and productivity than free cells.…”

Section: Enhancement Of C-19 Steroid Yield By Improving the Medium Co...mentioning

confidence: 99%

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New Insights into the Modification of the Non-Core Metabolic Pathway of Steroids in Mycolicibacterium and the Application of Fermentation Biotechnology in C-19 Steroid Production

Zhang

Xiao

Pan

et al. 2023

IJMS

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Androsta-4-ene-3,17-dione (AD), androsta-1,4-diene-3,17-dione (ADD), and 9α-hydroxy-4-androstene-3,17-dione (9-OHAD), which belong to C-19 steroids, are critical steroid-based drug intermediates. The biotransformation of phytosterols into C-19 steroids by Mycolicibacterium cell factories is the core step in the synthesis of steroid-based drugs. The production performance of engineered mycolicibacterial strains has been effectively enhanced by sterol core metabolic modification. In recent years, research on the non-core metabolic pathway of steroids (NCMS) in mycolicibacterial strains has made significant progress. This review discusses the molecular mechanisms and metabolic modifications of NCMS for accelerating sterol uptake, regulating coenzyme I balance, promoting propionyl-CoA metabolism, reducing reactive oxygen species, and regulating energy metabolism. In addition, the recent applications of biotechnology in steroid intermediate production are summarized and compared, and the future development trend of NCMS research is discussed. This review provides powerful theoretical support for metabolic regulation in the biotransformation of phytosterols.

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“…The overall process of the ADD bio-production from phytosterols in mycobacteria and mycolicibacteria has been unveiled, but some steps and mechanisms are still unknown (Liu et al 2018). Androsta-4-ene-3,17-dione (androstenedione, AD), sharing similar molecular structure and metabolic pathway, is usually produced as the byproduct in ADD-producing strains (Amin et al 2010; Zhang et al 2021;Zhao et al 2022). Additionally, the aqueous solubility of phytosterols is about 2 mg/L in aqueous media, limiting the e ciency of steroid bioconversion (Josefsen et al 2017).…”

Section: Introductionmentioning

confidence: 99%

Immobilization of rough morphotype Mycolicibacterium neoaurum R for androstadienedione production

Zhao,

Li,

et al. 2023

Biotechnol Lett

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Enhance androstadienedione (Androst-1,4-diene-3,17-dione, ADD) production of rough-type morphotype Mycolicibacterium neoaurum variant by repeated-batch fermentation of immobilized cells. ResultsM. neoaurum R was a rough colony morphotype variant, obtained from routine plating of smooth M. neoaurum strain CICC 21097. M. neoaurum R showed rougher cell surface and aggregated in broth. The ADD production of M. neoaurum R was notably lower than that of M. neoaurum CICC 21097 during the free cell fermentation, but the yield gap could be erased after proper cell immobilization. Subsequently, repeated-batch fermentation by immobilized M. neoaurum R was performed to shorten the production cycle and enhance the bio-production e ciency of ADD. Through the optimization of the immobilization carriers and the solvents for phytosterols, the ADD productivity of M. neoaurum R immobilized by semiexpanded perlite reached 0.075 g/L/h during the repeated-batch fermentation for 40 days. ConclusionsAlthough smooth strains that could homogenously suspended in broth seemed to be preferred in the steroid bioconversion, the rough-type strain M. neoaurum R might be able to nd their place by proper cell immobilization.

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Coexpression of VHb and MceG genes in Mycobacterium sp. Strain LZ2 enhances androstenone production via immobilized repeated batch fermentation

Cited by 14 publications

References 45 publications

Microbial metabolism of diosgenin by a novel isolated Mycolicibacterium sp. HK‐90: A promising biosynthetic platform to produce 19‐carbon and 21‐carbon steroids

Microbial metabolism of diosgenin by a novel isolated Mycolicibacterium sp. HK‐90: A promising biosynthetic platform to produce 19‐carbon and 21‐carbon steroids

New Insights into the Modification of the Non-Core Metabolic Pathway of Steroids in Mycolicibacterium and the Application of Fermentation Biotechnology in C-19 Steroid Production

Immobilization of rough morphotype Mycolicibacterium neoaurum R for androstadienedione production

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