A mutant form of 3-ketosteroid-Δ1-dehydrogenase gives altered androst-1,4-diene-3, 17-dione/androst-4-ene-3,17-dione molar ratios in steroid biotransformations by Mycobacterium neoaurum ST-095

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• 作者：Minglong Shao ; Xian Zhang ; Zhiming Rao…
• 关键词：Mycobacterium neoaurum ; Phytosterol ; 3 ; Ketosteroid ; Δ1 ; dehydrogenase ; Mutation
• 刊名：Journal of Industrial Microbiology and Biotechnology
• 出版年：2016
• 出版时间：May 2016
• 年：2016
• 卷：43
• 期：5
• 页码：691-701
• 全文大小：2,144 KB
• 参考文献：1.Anagnostopoulos C, Spizizen J (1961) Requirements for transformation in Bacillus subtilis. J Bacteriol 81:741PubMed PubMedCentral
  2.Bragin EY, Shtratnikova VY, Dovbnya D, Schelkunov M, Pekov YA, Malakho S, Egorova O, Ivashina T, Sokolov S, Ashapkin V (2013) Comparative analysis of genes encoding key steroid core oxidation enzymes in fast-growing Mycobacterium spp. strains. J Steroid Biochem Mol Biol 138:41–53CrossRef PubMed
  3.Choi K-P, Molnar I, Yamashita M, Murooka Y (1995) Purification and characterization of the 3-ketosteroid-Δ1-dehydrogenase of Arthrobacter simplex produced in Streptomyces lividans. J Biochem 117:1043–1049CrossRef PubMed
  4.Donova MV, Egorova OV (2012) Microbial steroid transformations: current state and prospects. Appl Microbiol Biotechnol 94:1423–1447. doi:10.​1007/​s00253-012-4078-0 CrossRef PubMed
  5.Fernandes P, Cruz A, Angelova B, Pinheiro HM, Cabral JMS (2003) Microbial conversion of steroid compounds: recent developments. Enzyme Microb Technol 32:688–705. doi:10.​1016/​S0141-0229(03)00029-2 CrossRef
  6.Fujii C, Morii S, Kadode M, Sawamoto S, Iwami M, Itagaki E (1999) Essential tyrosine residues in 3-Ketosteroid-Δ’-Dehydrogenase from Rhodococcus rhodochrous. J Biochem 126:662–667CrossRef PubMed
  7.Gordhan BG, Parish T (2001) Gene replacement using pretreated DNA. In: Parish T, Stoker NG (eds) Mycobacterium tuberculosis protocols, vol 54. Methods in Molecular Medicine. Humana Press Inc., New Jersey, pp 77–92. doi:10.​1385/​1-59259-147-7:​077
  8.Li Y, Lu F, Sun T, Du L (2007) Expression of ksdD gene encoding 3-ketosteroid-Δ1-dehydrogenase from Arthrobacter simplex in Bacillus subtilis. Lett Appl Microbiol 44:563–568CrossRef PubMed
  9.Liu C, Zhang X, Z-m Rao, Shao M-l WuD, Z-h Xu, Li H (2015) Mutation breeding of high 4-androstene-3, 17-dione-producing Mycobacterium neoaurum ZADF-4 by atmospheric and room temperature plasma treatment. J Zhejiang Univ Sci B 16:286–295CrossRef PubMed PubMedCentral
  10.Long S, Zhang X, Rao Z, Chen K, Xu M, Yang T, Yang S (2016) Amino acid residues adjacent to the catalytic cavity of tetramer l-asparaginase II contribute significantly to its catalytic efficiency and thermostability. Enzyme Microb Technol 82:15–22. doi:10.​1016/​j.​enzmictec.​2015.​08.​009 CrossRef PubMed
  11.Malaviya A, Gomes J (2008) Androstenedione production by biotransformation of phytosterols. Bioresour Technol 99:6725–6737. doi:10.​1016/​j.​biortech.​2008.​01.​039 CrossRef PubMed
  12.Marsheck WJ, Kraychy S, Muir RD (1972) Microbial degradation of sterols. Appl Microbiol 23:72–77PubMed PubMedCentral
  13.Martin CK (1977) Microbial cleavage of sterol side chains. Adv Appl Microbiol 22:29–58CrossRef PubMed
  14.Matsushita H, Itagaki E (1992) Essential histidine residue in 3-ketosteroid-Δ1-dehydrogenase. J Biochem 111:594–599PubMed
  15.Molnar I, Choi KP, Yamashita M, Murooka Y (1995) Molecular cloning, expression in Streptomyces livdans, 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 15:895–905CrossRef PubMed
  16.Morii S, Fujii C, Miyoshi T, Iwami M, Itagaki E (1998) 3-Ketosteroid-Δ1-dehydrogenase of Rhodococcus rhodochrous: sequencing of the genomic DNA and hyperexpression, purification, and characterization of the recombinant enzyme. J Biochem 124:1026–1032CrossRef PubMed
  17.Murray HC, Peterson DH (1952) Oxygenation of steroids by Mucorales fungi. Google Patents
  18.Plesiat P, Grandguillot M, Harayama S, Vragar S, Michel-Briand Y (1991) Cloning, sequencing, and expression of the Pseudomonas testosteroni gene encoding 3-oxosteroid delta 1-dehydrogenase. J Bacteriol 173:7219–7227PubMed PubMedCentral
  19.Rohman A, van Oosterwijk N, Thunnissen A-MW, Dijkstra BW (2013) Crystal structure and site-directed mutagenesis of 3-ketosteroid Δ1-dehydrogenase from Rhodococcus erythropolis SQ1 explain its catalytic mechanism. J Biol Chem 288:35559–35568CrossRef PubMed PubMedCentral
  20.Shao M, Rao Z, Zhang X, Xu M, Yang T, Li H, Xu Z, Yang S (2015) Bioconversion of cholesterol to 4-cholesten-3-one by recombinant Bacillus subtilis expressing choM gene encoding cholesterol oxidase from Mycobacterium neoaurum JC-12. J Chem Technol Biotechnol 90:1811–1820. doi:10.​1002/​jctb.​4491 CrossRef
  21.Shao M, Zhang X, Rao Z, Xu M, Yang T, Li H, Xu Z (2015) Enhanced production of Androst-1, 4-Diene-3, 17-Dione by Mycobacterium neoaurum JC-12 using three-stage fermentation strategy. PLoS One 10:e0137658CrossRef PubMed PubMedCentral
  22.Shao M, Zhang X, Rao Z, Xu M, Yang T, Li H, Xu Z, Yang S-T (2015) Efficient testosterone production by engineered Pichia pastoris co-expressing human 17β-hydroxysteroid dehydrogenase type 3 and Saccharomyces cerevisiae glucose 6-phosphate dehydrogenase with NADPH regeneration. Green Chem. doi:10.​1039/​C5GC02353J
  23.Shen YB, Wang M, Li HN, Wang YB, Luo JM (2012) Influence of hydroxypropyl-β-cyclodextrin on phytosterol biotransformation by different strains of Mycobacterium neoaurum. J Ind Microbiol Biotechnol 39:1253–1259CrossRef PubMed
  24.Van der Geize R, Hessels G, Van Gerwen R, Vrijbloed J, Van der Meijden P, Dijkhuizen L (2000) Targeted disruption of the kstD gene encoding a 3-Ketosteroid Δ1-Dehydrogenase isoenzyme of rhodococcus erythropolis strain SQ1. Appl Environ Microbiol 66:2029–2036CrossRef PubMedCentral
  25.van Oosterwijk N, Knol J, Dijkhuizen L, van der Geize R, Dijkstra BW (2012) Structure and catalytic mechanism of 3-ketosteroid-Δ4-(5α)-dehydrogenase from Rhodococcus jostii RHA1 genome. J Biol Chem 287:30975–30983CrossRef PubMed PubMedCentral
  26.Wang Z, Zhao F, Chen D, Li D (2006) Biotransformation of phytosterol to produce androsta-diene-dione by resting cells of Mycobacterium in cloud point system. Process Biochem 41:557–561CrossRef
  27.Wang ZF, Huang YL, Rathman JF, Yang ST (2002) Lecithin-enhanced biotransformation of cholesterol to androsta-1, 4-diene-3, 17-dione and androsta-4-ene-3, 17-dione. J Chem Technol Biotechnol 77:1349–1357CrossRef
  28.Wei W, Wang FQ, Fan SY, Wei DZ (2010) Inactivation and augmentation of the primary 3-ketosteroid-Δ1-dehydrogenase in Mycobacterium neoaurum NwIB-01: biotransformation of soybean phytosterols to 4-androstene-3,17-dione or 1,4-androstadiene-3,17-dione. Appl Environ Microbiol 76:4578–4582. doi:10.​1128/​aem.​00448-10 CrossRef PubMed PubMedCentral
  29.Xie RL, Shen YB, Qin N, Wang YB, Su LQ, Wang M (2015) Genetic differences in ksdD influence on the ADD/AD ratio of Mycobacterium neoaurum. J Ind Microbiol Biotechnol:1–7
  30.Xu XX, Jin FL, Yu XQ, Ji SX, Wang J, Cheng HX, Wang C, Zhang WQ (2007) Expression and purification of a recombinant antibacterial peptide, cecropin, from Escherichia coli. Protein Expr Purif 53:293–301CrossRef PubMed
  31.Xu Y-P, Qin J, Sun S-M, Liu T-T, Zhang X-L, Qian S-S, Zhu H-L (2014) Synthesis, crystal structures, molecular docking and urease inhibitory activity of nickel (II) complexes with 3-pyridinyl-4-amino-5-mercapto-1, 2, 4-triazole. Inorganica Chim Acta 423:469–476CrossRef
  32.Yao K, Xu L-Q, Wang F-Q, Wei D-Z (2014) Characterization and engineering of 3-ketosteroid-△1-dehydrogenase and 3-ketosteroid-9α-hydroxylase in Mycobacterium neoaurum ATCC 25795 to produce 9α-hydroxy-4-androstene-3,17-dione through the catabolism of sterols. Metab Eng 24:181–191. doi:10.​1016/​j.​ymben.​2014.​05.​005 CrossRef PubMed
  33.Zhang WQ, Shao ML, Rao ZM, Xu MJ, Zhang X, Yang TW, Li H, Xu ZH (2013) Bioconversion of 4-androstene-3,17-dione to androst-1,4-diene-3,17-dione by recombinant Bacillus subtilis expressing ksdd gene encoding 3-ketosteroid-Δ1-dehydrogenase from Mycobacterium neoaurum JC-12. J Steroid Biochem Mol Biol 135:36–42. doi:10.​1016/​j.​jsbmb.​2012.​12.​016 CrossRef PubMed
  34.Zhong C, Rao Z, Xia H, Xu Z, Fang H, Zhuge J (2009) Mutation breeding of Mycobacterium sp. for transformation of phytosterol into androst-1,4-diene-3,17-dione. Chem Bioeng 7:43–46 (in Chinese)
  
• 作者单位：Minglong Shao (1)
  Xian Zhang (1)
  Zhiming Rao (1)
  Meijuan Xu (1)
  Taowei Yang (1)
  Hui Li (2)
  Zhenghong Xu (2)
  Shangtian Yang (3)
  
  1. Laboratory of Applied Microorganisms and Metabolic Engineering, School of Biotechnology, Jiangnan University, 1800 Lihu Avenue, Wuxi, Jiangsu, 214122, People’s Republic of China
  2. Laboratory of Pharmaceutical Engineering, School of Pharmaceutical Sciences, Jiangnan University, Wuxi, Jiangsu, 214122, People’s Republic of China
  3. Department of Chemical and Biomolecular Engineering, The Ohio State University, Columbus, OH, 43210, USA
  
• 刊物类别：Biomedical and Life Sciences
• 刊物主题：Chemistry
  Biotechnology
  Genetic Engineering
  Biochemistry
  Bioinformatics
  Microbiology
  Microbial Genetics and Genomics
  
• 出版者：Springer Berlin / Heidelberg
• ISSN：1476-5535

文摘

Mycobacterium neoaurum ST-095 and its mutant M. neoaurum JC-12, capable of transforming phytosterol to androst-1,4-diene-3,17-dione (ADD) and androst-4-ene-3,17-dione (AD), produce very different molar ratios of ADD/AD. The distinct differences were related to the enzyme activity of 3-ketosteroid-Δ1-dehydrogenase (KSDD), which catalyzes the C_1,2 dehydrogenation of AD to ADD specifically. In this study, by analyzing the primary structure of KSDD_I (from M. neoaurum ST-095) and KSDD_II (from M. neoaurum JC-12), we found the only difference between KSDD_I and KSDD_II was the mutation of Val366 to Ser366. This mutation directly affected KSDD enzyme activity, and this result was confirmed by heterologous expression of these two enzymes in Bacillus subtilis. Assay of the purified recombinant enzymes showed that KSDD_II has a higher C_1,2 dehydrogenation activity than KSDD_I. The functional difference between KSDD_I and KSDD_II in phytosterol biotransformation was revealed by gene disruption and complementation. Phytosterol transformation results demonstrated that ksdd _I and ksdd _II gene disrupted strains showed similar ADD/AD molar ratios, while the ADD/AD molar ratios of the ksdd _I and ksdd _II complemented strains were restored to their original levels. These results proved that the different ADD/AD molar ratios of these two M. neoaurum strains were due to the differences in KSDD. Finally, KSDD structure analysis revealed that the Val366Ser mutation could possibly play an important role in stabilizing the active center and enhancing the interaction of AD and KSDD. This study provides a reliable theoretical basis for understanding the structure and catalytic mechanism of the Mycobacteria KSDD enzyme.