Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
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Effect of Polar Amino Acid Residue Substitution by Site-Directed Mutagenesis in the N-terminal Domain of Pseudomonas sp. Phytase on Enzyme Activity

  • Lee, Ga Hye (Department of Biotechnology, Pukyong National University) ;
  • Jang, Won Je (Department of Biotechnology, Pukyong National University) ;
  • Kim, Soyeong (Department of Biotechnology, Pukyong National University) ;
  • Kim, Yoonha (Department of Biotechnology, Pukyong National University) ;
  • Kong, In-Soo (Department of Biotechnology, Pukyong National University)
  • Received : 2020.03.11
  • Accepted : 2020.04.19
  • Published : 2020.07.28
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Abstract

The N-terminal domain of the Pseudomonas sp. FB15 phytase increases low-temperature activity and catalytic efficiency. In this study, the 3D structure of the N-terminal domain was predicted and substitutions for the amino acid residues of the region assumed to be the active site were made. The activity of mutants, in which alanine (A) was substituted for the original residue, was investigated at various temperatures and pH values. Significant differences in enzymatic activity were observed only in mutant E263A, suggesting that the amino acid residue at position 263 of the N-terminal domain is important in enzyme activity.

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References

  1. Kim OH, Kim YO, Shim JH, Jung YS, Jung WJ, Choi WC, et al. 2010. $\beta$-Propeller phytase hydrolyzes insoluble $Ca^{2+}$-phytate salts and completely abrogates the ability of phytate to chelate metal ions. Biochemistry 49: 10216-10227. https://doi.org/10.1021/bi1010249
  2. Kumar V, Sinha AK, Makkar HP, De Boeck G, Becker K. 2012. Phytate and phytase in fish nutrition. J. Anim. Physiol. Anim. Nutr. 96: 335-364. https://doi.org/10.1111/j.1439-0396.2011.01169.x
  3. Cao L, Wang W, Yang C, Yang Y, Diana J, Yakupitiyage A, et al. 2007. Application of microbial phytase in fish feed. Enzyme Microb. Tech. 40: 497-507. https://doi.org/10.1016/j.enzmictec.2007.01.007
  4. Lei XG, Porres JM, Mullaney EJ, Brinchpedersen H. 2007. Phytase: Source, Structure and Application, Industrial enzymes Section E: 505-529.
  5. Fu S, Sun J, Qian L, Li Z. 2008. Bacillus phytases: present scenario and future perspectives. Appl. Biochem. Biotechnol. 151: 1-8. https://doi.org/10.1007/s12010-008-8158-7
  6. Jang WJ, Lee JM, Park HD, Choi YB, Kong IS. 2018. N-terminal domain of the beta-propeller phytase of Pseudomonas sp. FB15 plays a role for retention of low-temperature activity and catalytic efficiency. Enzyme Microb. Technol. 117: 4-90.
  7. Jang WJ, Lee JM, Hasan MT, Kong IS. 2019. Fusion of the N-terminal domain of Pseudomonas sp. phytase with Bacillus sp. phytase and its effects on optimal temperature and catalytic efficiency. Enzyme Microb. Technol. 126: 69-76. https://doi.org/10.1016/j.enzmictec.2019.04.002
  8. He Z, Honeycutt CW. 2005. A modified molybdenum blue method for orthophosphate determination suitable for investigating enzymatic hydrolysis of organic phosphates. Commun Soil Sci. Plant Anal. 36: 1373-1383. https://doi.org/10.1081/CSS-200056954
  9. Tomschy A, Brugger R, Lehmann M, Svendsen A, Vogel K, Kostrewa D, et al. 2002. Engineering of phytase for improved activity at low pH. Appl. Environ. Microbiol. 68: 1907-1913. https://doi.org/10.1128/AEM.68.4.1907-1913.2002
  10. Tran TT, Mamo G, Búxo L, Le NN, Gaber Y, Mattiasson B, et al. 2011. Site-directed mutagenesis of an alkaline phytase: influencing specificity, activity and stability in acidic milieu. Enzyme Microb. Technol. 49: 177-182. https://doi.org/10.1016/j.enzmictec.2011.05.012
  11. Pace CN, Fu H, Fryar KL, Landua J, Trevino SR, Shirley BA, et al. 2011. Contribution of hydrophobic interactions to protein stability. J. Mol. Biol. 408: 514-528. https://doi.org/10.1016/j.jmb.2011.02.053
  12. Lee JM, Moon SY, Kim YR, Kim KW, Lee BJ, Kong IS. 2017. Improvement of thermostability and halostability of ${\beta}-1$, 3-1, 4-glucanase by substituting hydrophobic residue for Lys48. Int. J. Biol. Macromol. 94: 594-602. https://doi.org/10.1016/j.ijbiomac.2016.10.043