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Microbial sensor for new-generation cephalosporins based in a protein-engineered β- lactamase

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Abstract

A protein-engineered β-lactamase, constructed by site-directed mutagenesis inEscherichia coli (E104M/G238S), and having broadened specificity, was able to degrade cephalosporins of first, second, and third generations. Manipulations of culture conditions allowed an increase in β-lactamase specific activity by up to twofold. The resultant bacteria were used to construct an immersable whole-cell biosensor for the detection of new-generation cephalosporins. Cells were immobilized on agar membranes, which in turn were attached to the surface of a flat pH electrode, thus constituting a biosensor based on the detection of pH changes. The sensor was able to detect second- and third-generation cephalosporins: cefamandole (0.4-4 mM), cefotaxime (0.4-3.5 mM), and cefoperazone (0.3-1.85 mM). Response times were between 3.5 and 11 min, depending on the kind of cephalosporin tested. The biosensor was stable for at least 7 d, time during which up to 100 tests were performed.

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References

  1. Carlsen, M., Johansen, C., Min, R. W., Nielsen, J., Meier, H., and Lantreibecq, F. (1993),Anal. Chim. Acta 279, 51–58.

    Article  CAS  Google Scholar 

  2. Papariello, G. J., Mukherji, A. K., and Shearer, C. M. (1973),Anal Chem. 45, 790–792.

    Article  CAS  Google Scholar 

  3. Cullen, L. F., Rusling, J. F., Schleifer, A., and Papariello, G. J. (1974),Anal. Chem. 46, 1955–1961.

    Article  CAS  Google Scholar 

  4. Matsumoto, K., Seijo, H., Watanabe, T., Karube, I., Satoh, L., and Suzuki, S. (1979),Anal. Chim. Acta 105, 429–432.

    Article  CAS  Google Scholar 

  5. Caras, S. D., and Janata, J. (1980),Anal. Chem. 52, 1935–1937.

    Article  CAS  Google Scholar 

  6. Yerian, D. T., Christian, G. D., and Ruzicka, J. (1988),Anal. Chem. 60, 1250–1256.

    Article  CAS  Google Scholar 

  7. Gnanasekaran, R., and Mottola, H. A. (1985),Anal. Chem. 57, 1005–1009.

    Article  CAS  Google Scholar 

  8. Flanagan, M. T., and Carrol, N. J. (1986),Biotechnol. Bioeng. 28, 1093–1099.

    Article  CAS  Google Scholar 

  9. Tor, R., and Freeman, A. (1986),Anal. Chem. 58, 1042–1046.

    Article  CAS  Google Scholar 

  10. Olsson, B. (1988),Anal. Chim. Acta. 209, 123–133.

    Article  CAS  Google Scholar 

  11. Brand, U., Reinhardt, B., Rüther, F., Scheper, T., and Schügerl, K. (1989), InBiosensors: Applications in Medicine, Environmental Protedion and Process Control, GFB Monographs, Vol. 13, Scheller, E. F., and Schmidt, R. D., eds., VCH Verlagsgesellschaft, Weinheim, pp. 127–135.

    Google Scholar 

  12. Brand, U., Reinhardt, B., Rüther, F., Scheper, T., and Schügerl, K. (1990),Anal. Chim. Acta. 238, 201–210.

    Article  CAS  Google Scholar 

  13. Galindo, E., Bautista, D., García, J. L., and Quintero, R. (1990),Enzyme Microb. Technol. 12, 642–646.

    Article  CAS  Google Scholar 

  14. Meien, H., Kumaran, S., Danna, A. M., and Tran-Minh, C. (1991),Anal. Chim. Acta. 249, 405–411.

    Article  Google Scholar 

  15. Luo, S., and Walt, D. R. (1989),Anal. Chem. 61, 1069–1072.

    Article  CAS  Google Scholar 

  16. Kulp, T. J., Camins, J., Angel, S. M., Munkholm, C., and Walt, D. R. (1987),Anal. Chem. 59, 2849–2853.

    Article  CAS  Google Scholar 

  17. Carlyon, E. E., Lowe, C. R., Reid, D., and Bennion, I. (1992),Biosensors Bioeledronics 7, 141–146.

    Article  CAS  Google Scholar 

  18. Xie, X., Suleiman, A. A., and Guilbault, G. (1992),Biotechnol. Bioeng. 39, 1147–1150.

    Article  CAS  Google Scholar 

  19. Höbel, W., Papperger, A., and Polster, J. (1992),Biosensors Bioelectronics 7, 549–557.

    Article  Google Scholar 

  20. Decristoforo, G., and Danielsson, B. (1984),Anal. Chem. 56, 263–268.

    Article  CAS  Google Scholar 

  21. Caras, S. D., Janata, J., Saupe, D., and Schmidt, K. (1985),Anal Chem. 57, 1917–1925.

    Article  CAS  Google Scholar 

  22. Collatz, E., Labia, R., and Gutmann, L. (1990),Mol. Microbiol. 4, 1615–1620.

    Article  CAS  Google Scholar 

  23. Osuna, J., Viadiu, H., Fink, A. L., and Soberón, X. (1995),J. Biol. Chem. 270, 775–780.

    Article  CAS  Google Scholar 

  24. Viadiu, H., Osuna, J., Fink, A. L., and Soberón, X. (1995),J. Biol. Chem. 270, 781–787.

    Article  CAS  Google Scholar 

  25. Nicolini, C. (1995),Biosensors Bioelectronics 10, 105–127.

    Article  CAS  Google Scholar 

  26. Merino, E., Osuna, J., Bolivar, F., and Soberón, X. (1992),Biotechniques 12, 508–510.

    CAS  Google Scholar 

  27. Ross, W. G., and O’Callaghan, H. C. (1975),Methods Enzymol. 43, 69–85.

    CAS  Google Scholar 

  28. Melling, J. (1978), inTopics in Enzyme and Fermentation Biotechnology, vol. 2, Wiseman, A. ed., John Wiley, pp. 152–169.

  29. Richmond, M. (1975),Methods Enzymol. 43, 672–677.

    Article  CAS  Google Scholar 

  30. Rashtchian, A., Nouvarsani, R., Miller, G., and Gerlach, E. (1979),Antimicrob. Agents Chemother.16, 772–775.

    CAS  Google Scholar 

  31. Ryan, W., Parulekar, S. J., and Stark, B. C. (1989),Biotechnol. Bioeng. 34, 309–319.

    Article  CAS  Google Scholar 

  32. Ramirez, O. T., Flores, E., and Galindo, E. (1995),Asia-Pacific J. Mol. Biol. Biotechnol. 3, 165–197.

    Google Scholar 

  33. Galindo, E., Bolivar, F., and Quintero, R. (1990),J. Ferment. Bioeng. 69, 159–165.

    Article  CAS  Google Scholar 

  34. Van Der Schoot, B. H., and Bergveld, P. (1990),Anal. Chim. Acta 233, 49–57.

    Article  Google Scholar 

  35. Healey, B. G., and Walt, D. R. (1995),Anal. Chem. 67, 4471–4476.

    Article  CAS  Google Scholar 

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Authors and Affiliations

  1. Department of Bioengineering, Institute of Biotechnology, National University of Mexico, Apdo. Postal 510-3, 62250, Cuernavaca, Morelos, México

    Juan L. García, Carlos J. Nuñez, Edith G. González & Enrique Galindo

  2. Department of Molecular Recognition and Biostructure, Institute of Biotechnology, National University of Mexico, Apdo. Postal 510-3, 62250, Cuernavaca, Morelos, México

    Joel Osuna & Xavier Soberón

Authors
  1. Juan L. García
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  2. Carlos J. Nuñez
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  3. Edith G. González
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  4. Joel Osuna
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  5. Xavier Soberón
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  6. Enrique Galindo
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García, J.L., Nuñez, C.J., González, E.G. et al. Microbial sensor for new-generation cephalosporins based in a protein-engineered β- lactamase. Appl Biochem Biotechnol 73, 243–256 (1998). https://doi.org/10.1007/BF02785659

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  • DOI: https://doi.org/10.1007/BF02785659

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