Abstract
Traditionally, boldenone (BD), an important hormone drug and precursor for the synthesis of other steroids, was chemically produced. At present, the method of biotransformation is usually from androst-4-ene-3,17-dione (AD) to BD, but some bottlenecks seriously limit the wide industrial application of microbial transformation to produce BD. Recently, Pichia pastoris GS115 was found to be capable of transforming the C17-one to C17-alcohol of steroids, indicating that P. pastoris GS115 contain the gene for 17Hsd, but its 3-ketosteroid-Δ1-dehydrogenase has low activity. In this study, we successfully expressed a 3-ketosteroid-Δ1-dehydrogenase KsdD2 from Rhodococcus rhodochrous DSM43269 intracellularly in P. pastoris GS115. The engineered recombinant strains P. pastoris GS115 pPIC3.5K-ksdd2 was found to be capable of transforming AD to both androst-1,4-ene-3,17-dione (ADD) and BD. The transformation of AD to ADD and BD is certainly the result of synergism between exogenous KsdD2 and endogenous 17Hsd. In this way, BD could be obtained by biotransformation from AD, and the research developed in this study could provide theory basis for the wide industrial application of microbial transformation to produce BD.
Rui Tang and Peilin Ji contributed equally to this work.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Malaviya A, Gomes J (2008) Androstenedione production by biotransformation of phytosterols. Bioresour Technol 99(15):6725–6737
Brabander HF, Poelmans S, Schilt R et al (2004) Presence and metabolism of the anabolic steroid boldenone in various animal species: a review. Food Addit Contam 21(6):515–525
Fernandes P, Cabral JMS (2007) Phytosterols: applications and recovery methods. Bioresour Technol 98:2335–2350
Bhatti HN, Khera RA (2012) Biological transformations of steroidal compounds: a review. Steroids 77(12):1267–1290
Choi KP, Molnar IJ, Yamashita M (1995) Purification and characterization of the 3-ketosteroid-Δ1-dehydrogenase of Arthrobacter simplex produced in Streptomyces lividans. J Biochem 117(5):1043–1049
Chen MM, Wang FQ, Lin LC et al (2012) Characterization and application of fusidane antibiotic biosynethsis enzyme 3-ketosteroid-Δ1-dehydrogenase in steroid transformation. Appl Microbiol Biotechnol 96:133–142
Molnar I, Choi KP, Yamashita M et al (1995) Molecular cloning, expression in Streptomyces lividans, and analysis of a gene cluster from Arthrobacter simplex encoding 3-ketosteroid-Δ1-dehydrogenase, 3-ketosteroid-Δ5-isomerase and a hypothetical regulatory protein. Mol Microbiol 15:895–905
Morii S, Fujii C, Miyoshi T (1998) 3-Ketosteroid-Δ1-dehydrogenase of Rhodococcus rhodochrous sequencing of the genomic DNA and hyperexpression purification, and characterization of the recombinant enzyme. J Biochem 124:1026–1032
Liu Y, Shen YB, Qiao YQ et al (2016) The effect of 3-ketosteroid-Δ1-dehydrogenase isoenzymes on the transformation of AD to 9α-OH-AD by Rhodococcus rhodochrous DSM43269. J Ind Microbiol Biotechnol 43:1303–1311
Wagner M, Atrat PG, Wagner B et al (1992) Overexpression of a Rhodococcus erythropolis protein in Escherichia coli with immunological identity to the Rhodococcus steroid-1-dehydrogenase. Immunoelectron microscopic localization and electrophoretic studies. J Basic Microbiol 32:269–277
Wei W, Fan SY, Wang FQ et al (2014) Accumulation of androstadiene-dione by overexpression of heterologous 3-ketosteroid D1-dehydrogenase in Mycobacterium neoaurum NwIB-01. World J Microbiol Biotechnol 30:1947–1954
Shao ML, Zhang X, Rao ZM et al (2015) Enhanced production of androst-1, 4-Diene-3, 17-Dione by Mycobacterium neoaurum JC-12 using three-stage fermentation strategy. PLoS ONE 10(9):1–13
Xie RL, Shen YB, Qin N et al (2015) Genetic differences in ksdD influence on the ADD/AD ratio of Mycobacterium neoaurum. J Ind Microbiol Biotechnol 42:507–513
van der Geize R, Hessels GI, van Gerwen R et al (2002) Molecular and functional characterization of the kstD2 gene of Rhodococcus erythropolis SQ1 encoding a second 3-ketosteroid-Δ1-dehydrogenase isoenzyme. Microbiol 148:3285–3292
Singer Y, Shity H, Bar R (1991) Microbial transformation in a cyclodextrin medium. Part 2. reduction of androstenedione to testosterone by Saccharomyces cerevisiae. Appl Microbiol Biotechnol 35:731–737
Marina VD, Olga VE, Vera MN (2005) Steroid 17β-reduction by microorganisms–a review. Process Biochem 40(7):2253–2262
Wang M, Zhang LT, Shen YB et al (2009) Effects of hydroxypropyl-β-cyclodextrin on steroids 1-en-dehydrogenation biotransformation by Arthrobacter simplex TCCC 11037. J Mol Catal B Enzym 59:58–63
Zhang LT, Wang M, Shen YB et al (2009) Improvement of steroid biotransformation with hydroxypropyl-β-cyclodextrin induced complexation. Appl Biochem Biotechnol 159(3):642–654
Acknowledgements
This work was supported by National Natural Science Foundation of China (No. 21276196, 21406167), Key Project of Chinese Ministry of Education (213004A) and the Tianjin College students’ innovative entrepreneurial training plan (201410057106).
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2018 Springer Nature Singapore Pte Ltd.
About this paper
Cite this paper
Tang, R. et al. (2018). Biotransformation to Produce Boldenone by Pichia pastoris GS115 Engineered Recombinant Strains. In: Liu, H., Song, C., Ram, A. (eds) Advances in Applied Biotechnology. ICAB 2016. Lecture Notes in Electrical Engineering, vol 444. Springer, Singapore. https://doi.org/10.1007/978-981-10-4801-2_12
Download citation
DOI: https://doi.org/10.1007/978-981-10-4801-2_12
Published:
Publisher Name: Springer, Singapore
Print ISBN: 978-981-10-4800-5
Online ISBN: 978-981-10-4801-2
eBook Packages: Chemistry and Materials ScienceChemistry and Material Science (R0)