Advertisement

Utilization of one novel deep-sea microbial protease sin3406-1 in the preparation of ethyl (S)-3-hydroxybutyrate through kinetic resolution

  • Jinlong Huang
  • Yongkai Xu
  • Yun Zhang
  • Aijun Sun
  • Yunfeng Hu
Original Paper

Abstract

One novel protease sin3406-1 was identified from Streptomyces niveus SCSIO 3406, which was isolated from the deep sea of the South China Sea, and heterologously expressed in E. coli BL21(DE3). Protease sin3406-1 was further used as a green biocatalyst in the kinetic resolution of racemic ethyl-3-hydroxybutyrate. After careful process optimization, chiral product ethyl (S)-3-hydroxybutyrate was generated with an enantiomeric excess of over 99% and a conversion rate of up to 50% through direct hydrolysis of inexpensive racemic ethyl-3-hydroxybutyrate catalyzed by sin3406-1. Interestingly, protease sin3406-1 exhibited the same enantio-preference as that of esterase PHE21 during the asymmetric hydrolysis of the ester bonds of racemic ethyl-3-hydroxybutyrate. Through mutation studies and molecular docking, we also demonstrated that the four residues close to the catalytic center, S85, A86, Q87 and Y254, played key roles in both the hydrolytic activity and the enantioselectivity of protease sin3406-1, possibly through forming hydrogen bonds between the enzyme and the substrates. Deep-sea microbial proteases represented by sin3406-1 are new contributions to the biocatalyst library for the preparation of valuable chiral drug intermediates and chemicals through enzymatic kinetic resolution.

Keywords

Biocatalysis Marine microorganisms Novel protease Kinetic resolution Ethyl (S)-3-hydroxybutyrate 

Notes

Acknowledgements

We are grateful for the financial supports from Scientific and Technological Project of Ocean and Fishery from Guangdong Province (A201701C12), the Priority Research Program of the Chinese Academy of Sciences (XDA11030404) and Guangzhou Science and Technology Plan Projects (201510010012).

Compliance with ethical standards

Conflict of interest

The authors all declare that they have no conflict of interest.

Supplementary material

11274_2018_2513_MOESM1_ESM.docx (5.8 mb)
Supplementary material 1 (DOCX 5897 KB)

References

  1. Bhatt VS, Guan WY, Xue MY, Yuan HQ, Wang PG (2011) Insights into role of the hydrogen bond networks in substrate recognition by UDP-GaINAc 4-epimerases. Biochem Biophys Res Commun 412:232–237.  https://doi.org/10.1016/j.bbrc.2011.07.071 CrossRefPubMedPubMedCentralGoogle Scholar
  2. Brettin T, Davis JJ, Disz T, Edwards RA, Gerdes S, Olsen GJ et al (2015) RASTtk: a modular and extensible implementation of the RAST algorithm for building custom annotation pipelines and annotating batches of genomes. Sci Rep 5:8365.  https://doi.org/10.1038/srep08365 CrossRefPubMedPubMedCentralGoogle Scholar
  3. Bucciarelli M, Davoli P, Forni A, Moretti I, Prati F (1999) Enantioselective lipase-catalyzed acetylation of β-lactam precursors of carbapenem antibiotics. J Chem Soc Perkin Trans 1 1:2489–2494.  https://doi.org/10.1039/A903719E CrossRefGoogle Scholar
  4. Carnell AJ, Head R, Bassett D, Schneider M (2004) Efficient large scale stereoinversion of (R)-ethyl 3-hydroxybutyrate. Tetrahedron: Asymmetry 15:821–825.  https://doi.org/10.1016/j.tetasy.2003.12.005 CrossRefGoogle Scholar
  5. Castillo B, Delgado Y, Barletta G, Griebenow K (2010) Enantioselective transesterification catalysis by nanosized serine protease subtilisin Carlsberg particles in tetrahydrofuran. Tetrahedron 66:2175–2180.  https://doi.org/10.1016/j.tet.2010.01.053 CrossRefPubMedPubMedCentralGoogle Scholar
  6. Chen XL, Xie BB, Lu JT, He HL, Zhang YZ (2007) A novel type of subtilase from the psychrotolerant bacterium Pseudoalteromonas sp SM9913: catalytic and structural properties of deseasin MCP-01. Microbiology 153:2116–2125.  https://doi.org/10.1099/mic.0.2007/006056-0 CrossRefPubMedGoogle Scholar
  7. Deng J, Yao Z, Chen K, Yuan YA, Lin J, Wei D (2016) Towards the computational design and engineering of enzyme enantioselectivity: a case study by a carbonyl reductase from Gluconobacter oxydans. J Biotechnol 217:31–40.  https://doi.org/10.1016/j.jbiotec.2015.11.003 CrossRefPubMedGoogle Scholar
  8. Falus P, Cerioli L, Bajnóczi G, Boros Z, Weiser D, Nagy J et al (2016) A continuous-flow cascade reactor system for Subtilisin A- catalyzed dynamic kinetic resolution of N-tert-butyloxycarbonylphenylalanine ethyl thioester with benzylamine. Adv Synth Catal 358:1608–1617.  https://doi.org/10.1002/adsc.201500902 CrossRefGoogle Scholar
  9. Fillat A, Romea P, Pastor FIJ, Urpi F, Diaz P (2015) Kinetic resolution of esters from secondary and tertiary benzylic propargylic alcohols by an improved esterase-variant from Bacillus sp BP-7. Catal Today 255:16–20.  https://doi.org/10.1016/j.cattod.2014.12.041 CrossRefGoogle Scholar
  10. Fishman A, Eroshov M, Dee-Noor SS, van Mil J, Cogan U, Effenberger R (2001) A two-step enzymatic resolution process for large-scale production of (S)- and (R)-ethyl-3-hydroxybutyrate. Biotechnol Bioeng 74:256–263.  https://doi.org/10.1002/bit.1115 CrossRefPubMedGoogle Scholar
  11. Ghareib M, Fawzi EM, Aldossary NA (2014) Thermostable alkaline protease from Thermomyces lanuginosus: optimization, purification and characterization. Ann Microbiol 64:859–867.  https://doi.org/10.1007/s13213-013-0725-7 CrossRefGoogle Scholar
  12. Gu JL, Tong HF, Ye LD, Yu HW (2015) The role of aromatic residue W20 in the activity and enantioselectivity control of esterase BioH toward aryl substrate. Biochem Eng J 101:134–141.  https://doi.org/10.1016/j.bej.2015.05.004 CrossRefGoogle Scholar
  13. He J, Zhou L, Wang P, Zu L (2009) Microbial reduction of ethyl acetoacetate to ethyl (R)-3-hydroxybutyrate in an ionic liquid containing system. Process Biochem 44:316–321.  https://doi.org/10.1016/j.procbio.2008.11.007 CrossRefGoogle Scholar
  14. Huang J, Zhang Y, Hu Y (2016) Functional characterization of a marine bacillus esterase and its utilization in the stereo-selective production of D-methyl lactate. Appl Biochem Biotechnol 180:1467–1481.  https://doi.org/10.1007/s12010-016-2180-y CrossRefPubMedGoogle Scholar
  15. Iding H, Wirz B, Rogers-Evans M (2004) Chemo-enzymatic preparation of chiral 3-aminopyrrolidine derivatives. Tetrahedron 60:647–653.  https://doi.org/10.1016/j.tet.2003.10.118 CrossRefGoogle Scholar
  16. Li XJ, Zheng RC, Ma HY, Huang JF, Zheng YG (2014) Key residues responsible for enhancement of catalytic efficiency of Thermomyces lanuginosus lipase Lip revealed by complementary protein engineering strategy. J Biotechnol 188:29–35.  https://doi.org/10.1016/j.jbiotec.2014.08.004 CrossRefPubMedGoogle Scholar
  17. Li C, Teng X, Qi Y, Tang B, Shi H, Ma X et al (2016a) Conformational flexibility of a short loop near the active site of the SARS-3CLpro is essential to maintain catalytic activity. Sci Rep 6:20918.  https://doi.org/10.1038/srep20918 CrossRefPubMedPubMedCentralGoogle Scholar
  18. Li G, Yao P, Cong P, Ren J, Wang L, Feng J et al (2016b) New recombinant cyclohexylamine oxidase variants for deracemization of secondary amines by orthogonally assaying designed mutants with structurally diverse substrates. Sci Rep 6:24973.  https://doi.org/10.1038/srep24973 CrossRefPubMedPubMedCentralGoogle Scholar
  19. Liu Y, Tang T-X, Pei X-Q, Zhang C, Wu Z-L (2014) Identification of ketone reductase ChKRED20 from the genome of Chryseobacterium sp. CA49 for highly efficient anti-Prelog reduction of 3,5-bis(trifluoromethyl)acetophenone. J Mol Catal B 102:1–8.  https://doi.org/10.1016/j.molcatb.2014.01.009 CrossRefGoogle Scholar
  20. Long WS, Kow PC, Kamaruddin AH, Bhatia S (2005) Comparison of kinetic resolution between two racemic ibuprofen esters in an enzymic membrane reactor. Process Biochem 40:2417–2425.  https://doi.org/10.1016/j.procbio.2004.09.014 CrossRefGoogle Scholar
  21. Lylloff JE, Hansen LBS, Jepsen M, Sanggaard KW, Vester JK, Enghild JJ et al (2016) Genomic and exoproteomic analyses of cold- and alkaline-adapted bacteria reveal an abundance of secreted subtilisin-like proteases. Microb Biotechnol 9:245–256.  https://doi.org/10.1111/1751-7915.12343 CrossRefPubMedPubMedCentralGoogle Scholar
  22. Montanez-Clemente I, Alvira E, Macias M, Ferrer A, Fonceca M, Rodriguez J et al (2002) Enzyme activation in organic solvents: co-lyophilization of subtilisin Carlsberg with methyl-beta-cyclodextrin renders an enzyme catalyst more active than the cross-linked enzyme crystals. Biotechnol Bioeng 78:53–59.  https://doi.org/10.1002/bit.10182 CrossRefPubMedGoogle Scholar
  23. Morris GM, Huey R, Lindstrom W, Sanner MF, Belew RK, Goodsell DS et al (2009) AutoDock4 and AutoDockTools4: automated docking with selective receptor flexibility. J Comput Chem 30:2785–2791.  https://doi.org/10.1002/jcc.21256 CrossRefPubMedPubMedCentralGoogle Scholar
  24. Nakagawa A, Idogaki H, Kato K, Shinmyo A, Suzuki T (2006) Improvement on production of (R)-4-chloro-3-hydroxybutyrate and (S)-3-hydroxy-gamma-butyrolactone with recombinant Escherichia coli cells. J Biosci Bioeng 101:97–103.  https://doi.org/10.1263/jbb.101.97 CrossRefPubMedGoogle Scholar
  25. Paál TA, Liljeblad A, Kanerva LT, Forro E, Fulop F (2008) Directed (R)- or (S)-selective dynamic kinetic enzymatic hydrolysis of 1,2,3,4-tetrahydroisoquinoline-1-carboxylic esters. Eur J Org Chem.  https://doi.org/10.1002/ejoc.200800789 Google Scholar
  26. Petersen TN, Brunak S, von Heijne G, Nielsen H (2011) SignalP 4.0: discriminating signal peptides from transmembrane regions. Nat Methods 8:785–786.  https://doi.org/10.1038/nmeth.1701 CrossRefPubMedGoogle Scholar
  27. Song YX, Huang HB, Chen YC, Ding J, Zhang Y, Sun AJ et al (2013) Cytotoxic and antibacterial marfuraquinocins from the deep south China sea-derived Streptomyces niveus SCSIO 3406. J Nat Prod 76:2263–2268.  https://doi.org/10.1021/np4006025 CrossRefPubMedGoogle Scholar
  28. Sun Y, Cheng Q, Tian W, Wu X (2008) A substitutive substrate for measurements of β-ketoacyl reductases in two fatty acid synthase systems. J Biochem Biophys Methods 70:850–856.  https://doi.org/10.1016/j.jbbm.2007.10.005 CrossRefPubMedGoogle Scholar
  29. Tahir MN, Cho E, Mischnick P, Lee JY, Yu J-H, Jung S (2014) Pentynyl dextran as a support matrix for immobilization of serine protease subtilisin Carlsberg and its use for transesterification of N-acetyl-L-phenylalanine ethyl ester in organic media. Bioprocess Biosyst Eng 37:687–695.  https://doi.org/10.1007/s00449-013-1038-8 CrossRefPubMedGoogle Scholar
  30. Takaç S, Bakkal M (2007) Impressive effect of immobilization conditions on the catalytic activity and enantioselectivity of Candida rugosa lipase toward S-Naproxen production. Process Biochem 42:1021–1027.  https://doi.org/10.1016/j.procbio.2007.03.013 CrossRefGoogle Scholar
  31. Tessaro D, Cerioli L, Servi S, Viani F, D’Arrigo P (2011) L-Amino acid amides via dynamic kinetic resolution. Adv Synth Catal 353:2333–2338.  https://doi.org/10.1002/adsc.201100389 CrossRefGoogle Scholar
  32. Wang Y, Zhang Y, Hu Y (2016) Functional characterization of a robust marine microbial esterase and its utilization in the stereo-selective preparation of ethyl (S)-3-hydroxybutyrate. Appl Biochem Biotechnol 180:1–17.  https://doi.org/10.1007/s12010-016-2161-1 CrossRefGoogle Scholar
  33. Wei P, Gao J, Zheng G, Wu H, Zong M, Lou W (2016) Engineering of a novel carbonyl reductase with coenzyme regeneration in E. coli for efficient biosynthesis of enantiopure chiral alcohols. J Biotechnol 230:54–62.  https://doi.org/10.1016/j.jbiotec.2016.05.004 CrossRefPubMedGoogle Scholar
  34. Wrzosek K, García Rivera MA, Bettenbrock K, Seidel-Morgenstern A (2016) Racemization of undesired enantiomers: immobilization of mandelate racemase and application in a fixed bed reactor. Biotechnol J 11:453–463.  https://doi.org/10.1002/biot.201500494 CrossRefPubMedGoogle Scholar
  35. Yan Z, Li D, Yin X (2017) Review for chiral-at-metal complexes and metal-organic framework enantiomorphs. Sci Bull 62:1344–1354.  https://doi.org/10.1016/j.scib.2017.09.013 CrossRefGoogle Scholar
  36. Yu L, Xu Y, Wang X, Yu X (2007) Highly enantioselective hydrolysis of dl-menthyl acetate to l-menthol by whole-cell lipase from Burkholderia cepacia ATCC 25416. J Mol Catal B 47:149–154.  https://doi.org/10.1016/j.molcatb.2007.04.011 CrossRefGoogle Scholar
  37. Zheng G, Yu H, Zhang J, Xu J (2009) Enzymatic production of l-menthol by a high substrate concentration tolerable esterase from newly isolated Bacillus subtilis ECU0554. Adv Synth Catal 351:405–414.  https://doi.org/10.1002/adsc.200800412 CrossRefGoogle Scholar
  38. Zhu D, Yang Y, Majkowicz S, Pan TH-Y, Kantardjieff K, Hua L (2008) Inverting the enantioselectivity of a carbonyl reductase via substrate-enzyme docking-guided point mutation. Org Lett 10:525–528.  https://doi.org/10.1021/ol702638j CrossRefPubMedGoogle Scholar

Copyright information

© Springer Nature B.V. 2018

Authors and Affiliations

  • Jinlong Huang
    • 1
    • 2
    • 3
  • Yongkai Xu
    • 4
  • Yun Zhang
    • 1
    • 2
  • Aijun Sun
    • 1
    • 2
  • Yunfeng Hu
    • 1
    • 2
    • 5
  1. 1.CAS Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of OceanologyChinese Academy of SciencesGuangzhouPeople’s Republic of China
  2. 2.Guangdong Key Laboratory of Marine Materia Medica, South China Sea Institute of OceanologyChinese Academy of SciencesGuangzhouPeople’s Republic of China
  3. 3.College of Life ScienceGuangxi Normal UniversityGuilinPeople’s Republic of China
  4. 4.Affiliated Hospital of Shandong University of Traditional Chinese MedicineJinanPeople’s Republic of China
  5. 5.South China Sea Bio-Resource Exploitation and Utilization Collaborative Innovation CenterGuangzhouPeople’s Republic of China

Personalised recommendations