Applied Biochemistry and Biotechnology

, Volume 179, Issue 1, pp 75–93 | Cite as

Functional Characterization of a Novel Marine Microbial GDSL Lipase and Its Utilization in the Resolution of (±)-1-Phenylethanol

  • Dun Deng
  • Yun Zhang
  • Aijun Sun
  • Jiayuan Liang
  • Yunfeng HuEmail author


A novel GDSL lipase (MT6) was cloned from the genome of Marinactinospora thermotolerans SCSIO 00652 identified from the South China Sea. MT6 showed its maximum identity of 59 % with a putative lipase from Nocardiopsis dassonville. MT6 was heterologously expressed in E. coli BL21(DE3) and further functionally characterized. MT6 could efficiently resolve racemic 1-phenylethanol and generate (R)-1-phenylethanol with high enantiomeric excess (99 %) and conversion rate (54 %) through transesterification reactions after process optimization. Our report was the first one report about the utilization of one GDSL lipase in the preparation of chiral chemicals by transesterification reactions, and the optical selectivity of MT6 was interestingly opposite to those of other common lipases. GDSL lipases represented by MT6 possess great potential for the generation of valuable chiral chemicals in industry.


Novel GDSL lipase Kinetic resolution (R)-1-phenylethanol Opposite optical selectivity 



We are grateful for the financial supports from the National Natural Science Foundation of China (No. 21302199), Strategic Priority Research Program of the Chinese Academy of Sciences (XDA11030404), Project “Engineering High-Performance Microorganisms for Advanced Bio-Based Manufacturing” from the Chinese Academy of Sciences (KGZD-EW-606) and Guangzhou Science and Technology Plan Projects (201510010012). We also would like to thank for the constant helping from Professor Jianhua Ju and Professor Changsheng Zhang.


  1. 1.
    Suan, C. L., & Sarmidi, M. R. (2004). Immobilised lipase-catalysed resolution of (R, S)-1-phenylethanol in recirculated packed bed reactor. Journal of Molecular Catalysis B: Enzymatic, 28, 111–119.CrossRefGoogle Scholar
  2. 2.
    Schofer, S. H., Kaftzik, N., Wasserscheid, P., & Kragl, U. (2001). Enzyme catalysis in ionic liquids: lipase catalysed kinetic resolution of 1-phenylethanol with improved enantioselectivity. Chemical Communications, 5, 425–426.CrossRefGoogle Scholar
  3. 3.
    Frings, K., Koch, M., & Hartmeier, W. (1999). Kinetic resolution of 1-phenyl ethanol with high enantioselectivity with native and immobilized lipase in organic solvents. Enzyme and Microbial Technology, 25, 303–309.CrossRefGoogle Scholar
  4. 4.
    Dudzik, A., Snoch, W., Borowiecki, P., Opalinska-Piskorz, J., Witko, M., Heider, J., & Szaleniec, M. (2015). Asymmetric reduction of ketones and beta-keto esters by (S)-1-phenylethanol dehydrogenase from denitrifying bacterium Aromatoleum aromaticum. Applied Microbiology and Biotechnology, 99, 5055–5069.CrossRefGoogle Scholar
  5. 5.
    Hoffken, H. W., Duong, M., Friedrich, T., Breuer, M., Hauer, B., Reinhardt, R., Rabus, R., & Heider, J. (2006). Crystal structure and enzyme kinetics of the (S)-specific 1-phenylethanol dehydrogenase of the denitrifying bacterium strain EbN1. Biochemistry, 45, 82–93.CrossRefGoogle Scholar
  6. 6.
    Kniemeyer, O., & Heider, J. (2001). (S)-1-Phenylethanol dehydrogenase of Azoarcus sp strain EbN1, an enzyme of anaerobic ethylbenzene catabolism. Archives of Microbiology, 176, 129–135.CrossRefGoogle Scholar
  7. 7.
    Lozano, P., De Diego, T., Larnicol, M., Vaultier, M., & Iborra, J. L. (2006). Chemoenzymatic dynamic kinetic resolution of rac-1-phenylethanol in ionic liquids and ionic liquids/supercritical carbon dioxide systems. Biotechnology Letters, 28, 1559–1565.CrossRefGoogle Scholar
  8. 8.
    Li, X., Xu, L., Wang, G. L., Zhang, H. J., & Yan, Y. J. (2013). Conformation studies on Burkholderia cenocepacia lipase via resolution of racemic 1-phenylethanol in non-aqueous medium and its process optimization. Process Biochemistry, 48, 1905–1913.CrossRefGoogle Scholar
  9. 9.
    Cao, Y., Zhuang, Y., Yao, C. J., Wu, B., & He, B. F. (2012). Purification and characterization of an organic solvent-stable lipase from Pseudomonas stutzeri LC2-8 and its application for efficient resolution of (R, S)-1-phenylethanol. Biochemical Engineering Journal, 64, 55–60.CrossRefGoogle Scholar
  10. 10.
    Akoh, C. C., Lee, G. C., Liaw, Y. C., Huang, T. H., & Shaw, J. F. (2004). GDSL family of serine esterases/lipases. Progress in Lipid Research, 43, 534–552.CrossRefGoogle Scholar
  11. 11.
    Talker-Huiber, D., Jose, J., Glieder, A., Pressnig, M., Stubenrauch, G., & Schwab, H. (2003). Esterase EstE from Xanthomonas vesicatoria (Xv_EstE) is an outer membrane protein capable of hydrolyzing long-chain polar esters. Applied Microbiology and Biotechnology, 61, 479–487.CrossRefGoogle Scholar
  12. 12.
    Rathelot, J., Julien, R., Canioni, P., Coeroli, C., & Sarda, L. (1975). Studies on Effect of Bile-Salt and Colipase on Enzymatic Lipolysis - Improved Method for Determination of Pancreatic Lipase and Colipase. Biochimie, 57, 1117–1122.CrossRefGoogle Scholar
  13. 13.
    Chen, C. S., Fujimoto, Y., Girdaukas, G., & Sih, C. J. (1982). Quantitative-Analyses of Biochemical Kinetic Resolutions of Enantiomers. Journal of the American Chemical Society, 104, 7294–7299.CrossRefGoogle Scholar
  14. 14.
    Tian, X. P., Tang, S. K., Dong, J. D., Zhang, Y. Q., Xu, L. H., Zhang, S., & Li, W. J. (2009). Marinactinospora thermotolerans gen. nov., sp nov., a marine actinomycete isolated from a sediment in the northern South China Sea. International Journal of Systematic and Evolutionary Microbiology, 59, 948–952.CrossRefGoogle Scholar
  15. 15.
    Kumari, A., & Gupta, R. (2015). Functional characterization of a novel aspartic acid rich lipase, TALipC, from Trichosporon asahii MSR54: solvent-dependent enantio inversion during esterification of 1-phenylethanol. Biotechnology Letters, 37, 121–130.CrossRefGoogle Scholar
  16. 16.
    Cui, C. X., Xie, R., Tao, Y. F., Zeng, Q. Q., & Chen, B. Q. (2015). Improving performance of Yarrowia lipolytica lipase lip2-catalyzed kinetic resolution of (R, S)-1-phenylethanol by solvent engineering. Biocatalysis and Biotransformation, 33, 38–43.CrossRefGoogle Scholar
  17. 17.
    Li, X., Huang, S. S., Xu, L., & Yan, Y. J. (2013). Improving activity and enantioselectivity of lipase via immobilization on macroporous resin for resolution of racemic 1-phenylethanol in non-aqueous medium. BMC Biotechnology, 13, 986–991.Google Scholar
  18. 18.
    Xue, P., Yan, X. H., & Wang, Z. (2007). Lipase immobilized on HOOC-MCF: A highly enantioselective catalyst for transesterification resolution of (R, S)-1-phenylethanol. Chinese Chemical Letters, 18, 929–932.CrossRefGoogle Scholar
  19. 19.
    de los Rios, A. P., van Rantwijk, F., & Sheldon, R. A. (2012). Effective resolution of 1-phenyl ethanol by Candida antarctica lipase B catalysed acylation with vinyl acetate in protic ionic liquids (PILs). Green Chemistry, 14, 1584–1588.CrossRefGoogle Scholar
  20. 20.
    Petrovskaya, L. E., Novototskaya-Vlasova, K. A., Kryukova, E. A., Rivkina, E. M., Dolgikh, D. A., & Kirpichnikov, M. P. (2015). Cell surface display of cold-active esterase EstPc with the use of a new autotransporter from Psychrobacter cryohalolentis K5(T). Extremophiles, 19, 161–170.CrossRefGoogle Scholar
  21. 21.
    Prive, F., Kaderbhai, N. N., Girdwood, S., Worgan, H. J., Pinloche, E., Scollan, N. D., Huws, S. A., & Newbold, C. J. (2013). Identification and Characterization of Three Novel Lipases Belonging to Families II and V from Anaerovibrio lipolyticus 5ST. PloS One, 8, 1–9.CrossRefGoogle Scholar
  22. 22.
    Bae, S. Y., Ryu, B. H., Jang, E., Kim, S., & Kim, T. D. (2013). Characterization and immobilization of a novel SGNH hydrolase (Est24) from Sinorhizobium meliloti. Applied Microbiology and Biotechnology, 97, 1637–1647.CrossRefGoogle Scholar
  23. 23.
    Okamura, Y., Kimura, T., Yokouchi, H., Meneses-Osorio, M., Katoh, M., Matsunaga, T., & Takeyama, H. (2010). Isolation and Characterization of a GDSL Esterase from the Metagenome of a Marine Sponge-associated Bacteria. Marine Biotechnology, 12, 395–402.CrossRefGoogle Scholar
  24. 24.
    Ding, J. M., Yu, T. T., Liang, L. M., Xie, Z. R., Yang, Y. J., Zhou, J. P., Xu, B., Li, J. J., & Huang, Z. X. (2014). Biochemical Characterization of a GDSL-Motif Esterase from Bacillus sp K91 with a New Putative Catalytic Mechanism. Journal of Microbiology and Biotechnology, 24, 1551–1558.CrossRefGoogle Scholar
  25. 25.
    Yang, Z. X., Zhang, Y., Shen, T. T., Xie, Y., Mao, Y. M., & Ji, C. N. (2013). Cloning, expression and biochemical characterization of a novel, moderately thermostable GDSL family esterase from Geobacillus thermodenitrificans T2. Journal of Bioscience and Bioengineering, 115, 133–137.CrossRefGoogle Scholar
  26. 26.
    Feller, G., Zekhnini, Z., LamotteBrasseur, J., & Gerday, C. (1997). Enzymes from cold-adapted microorganisms - The class C beta-lactamase from the Antarctic psychrophile Psychrobacter immobilis A5. European Journal of Biochemistry, 244, 186–191.CrossRefGoogle Scholar
  27. 27.
    Russell, R. J. M., Gerike, U., Danson, M. J., Hough, D. W., & Taylor, G. L. (1998). Structural adaptations of the cold-active citrate synthase from an Antarctic bacterium. Structure, 6, 351–361.CrossRefGoogle Scholar
  28. 28.
    Belcastro, M., Marino, T., Russo, N., & Toscano, M. (2005). Interaction of cysteine with Cu2+ and Group IIb (Zn2+, Cd2+, Hg2+) metal cations: a theoretical study. Journal of Mass Spectrometry, 40, 300–306.CrossRefGoogle Scholar
  29. 29.
    Cote, A., & Shareck, F. (2008). Cloning, purification and characterization of two lipases from Streptomyces coelicolor A3(2). Enzyme and Microbial Technology, 42(5), 381–388.CrossRefGoogle Scholar
  30. 30.
    Mander, P., Cho, S. S., Simkhada, J. R., Choi, Y. H., Park, D. J., & Yoo, J. C. (2012). An organic solvent-tolerant lipase from Streptomyces sp CS133 for enzymatic transesterification of vegetable oils in organic media. Process Biochemistry, 47, 635–642.CrossRefGoogle Scholar
  31. 31.
    Farrokh, P., Yakhchali, B., & Karkhane, A. A. (2014). Cloning and characterization of newly isolated lipase from Enterobacter sp. Bn12. Brazilian Journal of Microbiology, 45, 677–687.CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  • Dun Deng
    • 1
    • 2
    • 3
  • Yun Zhang
    • 1
    • 3
  • Aijun Sun
    • 1
    • 3
  • Jiayuan Liang
    • 1
    • 3
  • Yunfeng Hu
    • 1
    • 3
    • 4
    Email author
  1. 1.Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of OceanologyChinese Academy of SciencesGuangzhouPeople’s Republic of China
  2. 2.University of Chinese Academy of SciencesBeijingPeople’s Republic of China
  3. 3.Guangdong Key Laboratory of Marine Materia Medica, South China Sea Institute of OceanologyChinese Academy of SciencesGuangzhouPeople’s Republic of China
  4. 4.South China Sea Bio-Resource Exploitation and Utilization Collaborative Innovation CenterGuangzhouPeople’s Republic of China

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