Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
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Gene Cloning, Expression, and Characterization of a New Carboxylesterase from Serratia sp. SES-01: Comparison with Escherichia coli BioHe Enzyme

  • Kwon, Min-A (Chemical Biotechnology Research Center, Korea Research Institute of Chemical Technology) ;
  • Kim, Hyun-Suk (Chemical Biotechnology Research Center, Korea Research Institute of Chemical Technology) ;
  • Oh, Joon-Young (Taegwang Chemicals Co. Ltd.) ;
  • Song, Bong-Keun (Chemical Biotechnology Research Center, Korea Research Institute of Chemical Technology) ;
  • Song, Jae-Kwang (Chemical Biotechnology Research Center, Korea Research Institute of Chemical Technology)
  • Published : 2009.02.28
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Abstract

The carboxylesterase-encoding gene(bioHs) of a newly isolated strain, Serratia sp. SES-01, was cloned from the genomic DNA library by detecting formation of transparent halo around the colony on LB-tributyrin agar plates. The amino acid sequence of BioHs was highly similar to the members of the BioH enzyme family involved in the biotin biosynthetic pathway; it showed the highest similarity(91%) with that of Serratia proteamaculans. To compare BioHs with other BioH enzymes, the relatively well-known bioHe gene of E. coli was cloned with PCR. After we achieved high-level expression of soluble BioHs and BioHe through the exploration of different culture conditions, the purified BioHs and BioHe enzymes were characterized in terms of specificity, activity, and stability. BioHe was generally more robust to a change in temperature and pH and an addition of organic solvents than BioHs. The two enzymes exhibited a strong preference for carboxylesterase rather than for thioesterase and were optimal at relatively low temperatures($20-40^{\circ}C$) and alkaline pHs(7.5-9.0). The results in this study strongly suggested that both the BioHs and BioHe enzymes would be potential candidates for use as a carboxylesterase in many industrial applications.

Keywords

References

  1. Akatsuka, H., E. Kawai, N. Sakurai, and K. Omori. 2003. The Serratia marcescens bioH gene encodes an esterase. Gene 302: 185-192 https://doi.org/10.1016/S0378111902011502
  2. Bornscheuer, U. T. 2002. Microbial carboxyl esterases: Classification, properties and application in biocatalysis. FEMS Microbiol. Rev. 26: 73-81 https://doi.org/10.1111/j.1574-6976.2002.tb00599.x
  3. Bower, S., J. B. Perkins, R. R. Yocum, C. L. Howitt, P. Rahaim, and J. Pero. 1996. Cloning, sequencing, and characterization of the Bacillus subtilis biotin biosynthetic operon. J. Bacteriol. 178: 4122-4130
  4. Eom, G. T., J. S. Rhee, and J. K. Song. 2006. An efficient secretion system for type I secretion pathway-dependent lipase, TliA, in Escherichia coli: Effect of relative expression levels and expression timing of passenger protein and ABC transporter. J. Microbiol. Biotechnol. 16: 1422-1428
  5. Ifuku, O., H. Miyaoka, N. Koga, J. Kishimoto, S. Haze, Y. Wachi, and M. Kajiwara. 1994. Origin of carbon atoms of biotin. 13CNMR studies on biotin biosynthesis in Escherichia coli. Eur. J. Biochem. 220: 585-591 https://doi.org/10.1111/j.1432-1033.1994.tb18659.x
  6. Lemoine, Y., A. Wach, and J. M. Jeltsch. 1996. To be free or not: The fate of pimelate in Bacillus sphaericus and in Escherichia coli. Mol. Microbiol. 19: 645-647 https://doi.org/10.1046/j.1365-2958.1996.t01-4-442924.x
  7. Ploux, O., P. Soularue, A. Marquet, R. Gloeckler, and Y. Lemoine. 1992. Investigations of the first step of biotin biosynthesis in Bacillus sphaericus: Purification and characterization of the pimeloyl-CoA synthetase, and uptake of pimelate. Biochem. J. 287: 685-690 https://doi.org/10.1042/bj2870685
  8. Rodionov, D. A., A. A. Mironov, and M. S. Gelfand. 2002. Conservation of the biotin regulon and the BirA regulatory signal in Eubacteria and Archaea. Genome Res. 12: 1507-1516 https://doi.org/10.1101/gr.314502
  9. Sakurai, N., Y. Imai, and S. Komatsubara. 1995. Instability of the mutated biotin operon plasmid in a biotin-producing mutant of Serratia marcescens. J. Biotechnol. 43: 11-19 https://doi.org/10.1016/0168-1656(95)00103-9
  10. Sakurai, N., Y. Imai, M. Masuda, S. Komatsubara, and T. Tosa. 1994. Improvement of a d-biotin-hyperproducing recombinant strain of Serratia marcescens. J. Biotechnol. 36: 63-73 https://doi.org/10.1016/0168-1656(94)90024-8
  11. Sanishvili, R., A. F. Yakunin, R. A. Laskowski, T. Skarina, E. Evdokimova, A. Doherty-Kirby, et al. 2003. Integrating structure, bioinformatics, and enzymology to discover function: BioH, a new carboxylesterase from Escherichia coli. J. Biol. Chem. 278: 26039-26045 https://doi.org/10.1074/jbc.M303867200
  12. Sproer, C., U. Mendrock, J. Swiderski, E. Lang, and E. Stackebrandt. 1999. The phylogenetic position of Serratia, Buttiauxella and some other genera of the family Enterobacteriaceae. Int. J. Syst. Bacteriol. 49: 1433-1438 https://doi.org/10.1099/00207713-49-4-1433
  13. Tomczyk, N. H., J. E. Nettleship, R. L. Baxter, H. J. Crichton, S. P. Webster, and D. J. Campopiano. 2002. Purification and characterisation of the BIOH protein from the biotin biosynthetic pathway. FEBS Lett. 513: 299-304 https://doi.org/10.1016/S0014-5793(02)02342-6
  14. Watanabe, H., T. Shiratori, H. Shoji, S. Miyatake, Y. Okazaki, K. Ikuta, T. Sato, and T. Saito. 1997. A novel acyl-CoA thioesterase enhances its enzymatic activity by direct binding with HIV Nef. Biochem. Biophys. Res. Commun. 238: 234-239 https://doi.org/10.1006/bbrc.1997.7217
  15. Webb, M. E., A. Marquet, R. R. Mendel, F. Rébeillé, and A. G. Smith. 2007. Elucidating biosynthetic pathways for vitamins and cofactors. Nat. Prod. Rep. 24: 988-1008 https://doi.org/10.1039/b703105j
  16. Xie, X., W. W. Wong, and Y. Tang. 2007. Improving simvastatin bioconversion in Escherichia coli by deletion of bioH. Metab. Eng. 9: 379-386 https://doi.org/10.1016/j.ymben.2007.05.006