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Directed Evolution of a β-Glucosidase for Improved Functions as a Reporter in Protein Expression

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Precisely reliable and quantitative reporters can provide phenotypes that are consistent with research goals in protein expression. Here, we developed an improved reporter mATglu III 5 by directed evolution using a versatile β-glucosidase ATglu derived from Agrobacterium tumefaciens. When expressed in hosts, a vector containing this mutant distinctly showed a colored or fluorescent phenotype, according to the supplemented substrate, without any inducer. Analysis of mATglu III 5 showed it to be fully functional in fusion state with oligomeric proteins, especially under non-induction conditions, thereby offering an alternative to conventional reporters.

Abstract

Precisely reliable and quantitative reporters can provide phenotypes that are consistent with research goals in protein expression. Here, we developed an improved reporter mATglu III 5 by directed evolution using a versatile β-glucosidase ATglu derived from Agrobacterium tumefaciens. When expressed in hosts, a vector containing this mutant distinctly showed a colored or fluorescent phenotype, according to the supplemented substrate, without any inducer. Analysis of mATglu III 5 showed it to be fully functional in fusion state with oligomeric proteins, especially under non-induction conditions, thereby offering an alternative to conventional reporters.

참고문헌 (15)

  1. Naylor LH, "Reporter gene technology: the future looks bright", Biochem. Pharmacol, 58, pp.749~757 (1999)
  2. Jiang T, "Recent developments of biological reporter technology for detecting gene expression", Biotechnol. Genet. Eng. Rev, 25, pp.41~75 (2008)
  3. Tsien RY, "The green fluorescent protein", Annu. Rev. Biochem, 67, pp.509~544 (1998)
  4. Pédelacq JD, "Engineering and characterization of a superfolder green fluorescent protein", Nat. Biotechnol, 24, pp.79~88 (2006)
  5. Shaner NC, "Improved monomeric red, orange and yellow fluorescent proteins derived from Discosoma sp. red fluorescent protein", Nat. Biotechnol, 22, pp.1567~1572 (2004)
  6. Shuman HA, "The art and design of genetic screens : Escherichia coli", Nat. Rev. Genet, 4, pp.419~431 (2003)
  7. Iost I, "The stability of Escherichia coli lacZ mRNA depends upon the simultaneity of its synthesis and translation", EMBO J, 14, pp.3252~3261 (1995)
  8. Wu N, "Bacterial expression and reengineering of Gaussia princeps luciferase and its use as a reporter protein", Biochim. Biophys. Acta, 1854, pp.1392~1399 (2015)
  9. Chernov KG, "Nearinfrared fluorescent proteins, biosensors, and optogenetic tools engineered from phytochromes", Chem. Rev, 117, pp.6423~6446 (2017)
  10. Kim JY, "Mining and identification of a glucosidase family enzyme with high activity toward the plant extract indican", J. Mol. Catal. B, 57, pp.284~291 (2009)
  11. Kim JY, "Characterization of an indican-hydrolyzing enzyme from Sinorhizobium meliloti", Process Biochem, 45, pp.892~896 (2010)
  12. Cheong DE, "A cloning vector employing a versatile β-glucosidase as an indicator for recombinant clones", Anal. Biochem, 425, pp.166~168 (2012)
  13. Han SS, "Antisense RNA-based high-throughput screen system for directed evolution of quorum quenching enzymes", ACS Chem. Biol, 10, pp.2598~2605 (2015)
  14. Park WJ, "Over-expression of recombinant proteins with N-terminal His-tag via subcellular uneven distribution in Escherichia coli", Acta Biochim. Biophys. Sin, 47, pp.488~495 (2015)
  15. Choi ES, "Generation of a fast maturating red fluorescent protein by a combined approach of elongation mutagenesis and functional salvage screening", Biochem. Biophys. Res. Commun, 391, pp.598~603 (2010)

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