Hostname: page-component-586b7cd67f-gb8f7 Total loading time: 0 Render date: 2024-11-24T17:25:30.421Z Has data issue: false hasContentIssue false
  • English
  • Français

Purified proteases of two Antarctic bacteria: from screening to characterization

Published online by Cambridge University Press:  20 October 2021

Christian Peralta-Figueroa Open the ORCID record for Christian Peralta-Figueroa [Opens in a new window] ,
José Martínez-Oyanedel ,
Marta Bunster  and
Gerardo González-Rocha Open the ORCID record for Gerardo González-Rocha [Opens in a new window]
Christian Peralta-Figueroa
Affiliation:
Laboratorio de Investigación en Agentes Antibacterianos (LIAA), Departamento de Microbiología, Facultad de Ciencias Biológicas, Universidad de Concepción, Concepción, 4070386, Chile Laboratorio de Biofísica Molecular, Departamento de Bioquímica y Biología Molecular, Facultad de Ciencias Biológicas, Universidad de Concepción, Concepción, 4070386, Chile Ceibo. Av. Quilín 4757, Macul, Santiago de Chile, 7811013, Chile
José Martínez-Oyanedel
Affiliation:
Laboratorio de Biofísica Molecular, Departamento de Bioquímica y Biología Molecular, Facultad de Ciencias Biológicas, Universidad de Concepción, Concepción, 4070386, Chile
Marta Bunster
Affiliation:
Laboratorio de Biofísica Molecular, Departamento de Bioquímica y Biología Molecular, Facultad de Ciencias Biológicas, Universidad de Concepción, Concepción, 4070386, Chile
Gerardo González-Rocha*
Affiliation:
Laboratorio de Investigación en Agentes Antibacterianos (LIAA), Departamento de Microbiología, Facultad de Ciencias Biológicas, Universidad de Concepción, Concepción, 4070386, Chile Programa Especial de Ciencia Antártica y Subantártica (PCAS), Universidad de Concepción, Concepción, 4070386, Chile
*
[email protected]
Get access Rights & Permissions [Opens in a new window]

Abstract

Proteases are widely used in industrial processes, and the discovery of new, more kinetically efficient proteases can have a positive impact on industry. Enzymes from Antarctic microorganisms exhibit cold-adaptive properties, making them useful in biotechnology. The cold and harsh environment of Antarctica makes it a valuable source for new biotechnologically related enzymes. In this study, we characterized two cold-adapted proteases purified from Pseudoalteromonas issachenkonii P14M1-4 and Flavobacterium frigidimaris ANT34-7, isolated from King George Island, Antarctica, and compared these with proteases from the non-cold-adapted bacteria Bacillus licheniformis and Geobacillus stearothermophilus. The best temperature growing conditions were used for protease purification and characterization. The protease from P. issachenkonii P14M1-4 was identified as a 40–43 kDa metal-dependent subtilisin-like serine protease and the protease from F. frigidimaris ANT34-7 was identified as a 28 kDa metalloprotease. The enzymes showed an optimum temperature of between 35°C and 40°C and an optimum pH in the neutral to alkaline range. Their activation energies, catalytic constants and growth capacities at different temperatures categorize them as cold-adapted enzymes. We conclude that the characteristics exhibited by these proteases make them useful for biotechnological purposes requiring high activity at low temperatures. Moreover, to the best of our knowledge, this is the first characterization of a cold-adapted protease from F. frigidimaris.

Keywords

Antarcticacold-adapted proteasesenzyme adaptationenzymeskinetic parametersprotein purification
Type
Biological Sciences
Information
Antarctic Science , Volume 33 , Issue 6 , December 2021 , pp. 633 - 644
Check for updates
Copyright
Copyright © Antarctic Science Ltd 2021

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

Al-Ghanayem, A.A. & Joseph, B. 2020. Current prospective in using cold-active enzymes as eco-friendly detergent additive. Applied Microbiology and Biotechnology, 104, 2871–2882.CrossRefGoogle ScholarPubMed
Alexander, P.A., Ruan, B. & Bryan, P.N. 2001. Cation-dependent stability of subtilisin. Biochemistry, 40, 10634–10639.CrossRefGoogle ScholarPubMed
Bradford, M.M. 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry, 72, 248–254.CrossRefGoogle ScholarPubMed
Bruno, S., Coppola, D., di Prisco, G., Giordano, D. & Verde, C. 2019. Enzymes from marine polar regions and their biotechnological applications. Marine Drugs, 17, 544.CrossRefGoogle ScholarPubMed
Cavicchioli, R., Charlton, T., Ertan, H., Omar, S.M., Siddiqui, K.S. & Williams, T.J. 2011. Biotechnological uses of enzymes from psychrophiles. Microbial Biotechnology, 4, 449–460.CrossRefGoogle ScholarPubMed
Ceruso, M., Howe, N. & Malthouse, J.P.G. 2012. Mechanism of the binding of Z-L-tryptophan and Z-L-phenylalanine to thermolysin and stromelysin-1 in aqueous solutions. Biochimica et Biophysica Acta, 1824, 303–310.CrossRefGoogle ScholarPubMed
Dong, Z., Chen, Z., Wang, H., Tian, K., Jin, P., Liu, X., et al. (2017). Tandem mass tag-based quantitative proteomics analyses reveal the response of Bacillus licheniformis to high growth temperatures. Annals of Microbiology, 67, 501–510.CrossRefGoogle Scholar
Eisenthal, R., Danson, M.J. & Hough, D.W. 2007. Catalytic efficiency and k cat/KM: a useful comparator? Trends in Biotechnology, 25, 247–249.CrossRefGoogle ScholarPubMed
Feller, G. 2013. Psychrophilic enzymes: from folding to function and biotechnology. Scientifica, 2013, 512840.CrossRefGoogle ScholarPubMed
Furhan, J. 2020. Adaptation, production, and biotechnological potential of cold-adapted proteases from psychrophiles and psychrotrophs: recent overview. Journal of Genetic Engineering and Biotechnology, 18, 1–13.CrossRefGoogle ScholarPubMed
Gerday, C. 2013. Psychrophily and catalysis. Biology, 2, 719–741.CrossRefGoogle ScholarPubMed
González-Rocha, G., Muñoz-Cartes, G., Canales-Aguirre, C.B., Lima, C.A., Domínguez-Yévenes, M., Bello-Toledo, H. & Hernández, C.E. 2017. Diversity structure of culturable bacteria isolated from the Fildes Peninsula (King George Island, Antarctica): a phylogenetic analysis perspective. PLoS ONE, 12, e0179390.CrossRefGoogle Scholar
Heussen, C. & Dowdle, E.B. 1980. Electrophoretic analysis of plasminogen activators in polyacrylamide gels containing sodium dodecyl sulfate and copolymerized substrates. Analytical Biochemistry, 102, 196–202.CrossRefGoogle ScholarPubMed
Ivanova, E.P., Sawabe, T., Alexeeva, Y.V., Lysenko, A.M., Gorshkova, N.M., Hayashi, K., et al. 2002. Pseudoalteromonas issachenkonii sp. nov., a bacterium that degrades the thallus of the brown alga Fucus evanescens. International Journal of Systematic and Evolutionary Microbiology, 52, 229–234.CrossRefGoogle ScholarPubMed
Joshi, S. & Satyanarayana, T. 2013. Biotechnology of cold-active proteases. Biology (Basel), 2, 755–783.Google ScholarPubMed
Kakagianni, M., Gougouli, M. & Koutsoumanis, K.P. 2016. Development and application of Geobacillus stearothermophilus growth model for predicting spoilage of evaporated milk. Food Microbiology, 57, 28–35.CrossRefGoogle ScholarPubMed
Krishnan, A., Convey, P., Gonzalez-Rocha, G. & Alias, S.A. 2016. Production of extracellular hydrolase enzymes by fungi from King George Island. Polar Biology, 39, 65–76.CrossRefGoogle Scholar
Kulakova, L., Galkin, A., Kurihara, T., Yoshimura, T. & Esaki, N. 1999. Cold-active serine alkaline protease from the psychrotrophic bacterium Shewanella strain Ac10: gene cloning and enzyme purification and characterization. Applied and Environmental Microbiology, 65, 611–617.CrossRefGoogle ScholarPubMed
Laemmli, U.K. 1970. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature, 227, 680–685.CrossRefGoogle ScholarPubMed
Liu, J., Liu, W., Xing, S., Zhang, X., He, H., Chen, J., et al. 2021. Diversity of protease-producing bacteria in the soils of the South Shetland Islands, Antarctica. Antonie van Leeuwenhoek, 114, 457–464.CrossRefGoogle ScholarPubMed
Lonhienne, T., Gerday, C. & Feller, G. 2000. Psychrophilic enzymes: revisiting the thermodynamic parameters of activation may explain local flexibility. Biochimica et Biophysica Acta, 1543, 1–10.CrossRefGoogle ScholarPubMed
Luo, M., Eaton, C.N., Hess, K.R., Phillips-Piro, C.M., Brewer, S.H. & Fenlon, E.E. 2019. Paired spectroscopic and crystallographic studies of proteases. ChemistrySelect, 4, 9836.CrossRefGoogle ScholarPubMed
Martínez-Rosales, C. & Castro-Sowinski, S. 2011. Antarctic bacterial isolates that produce cold-active extracellular proteases at low temperature but are active and stable at high temperature. Polar Research, 30, 7123.CrossRefGoogle Scholar
McMahon, G.P. 2005. Mass spectrometry | peptides and proteins. In Worsfold, P., Poole, C., Townshend, A., & Miró, M., eds., Encyclopedia of analytical science. Amsterdam: Elsevier, 501–509.CrossRefGoogle Scholar
Millet, J. 1970. Characterization of proteinases excreted by Bacillus subtilis Marburg strain during sporulation. Journal of Applied Bacteriology, 33, 207–219.CrossRefGoogle ScholarPubMed
Nogi, Y., Soda, K. & Oikawa, T. 2005. Flavobacterium frigidimaris sp. nov., isolated from Antarctic seawater. Systematic and Applied Microbiology, 28, 310–315.CrossRefGoogle ScholarPubMed
Park, H.J., Lee, C.W., Kim, D., Do, H., Han, S.J., Kim, J.E., et al. 2018. Crystal structure of a cold-active protease (Pro21717) from the psychrophilic bacterium, Pseudoalteromonas arctica PAMC 21717, at 1.4 Å resolution: Structural adaptations to cold and functional analysis of a laundry detergent enzyme. PLoS ONE, 13, e0191740.CrossRefGoogle ScholarPubMed
Pelmenschikov, V., Blomberg, M.R. & Siegbahn, P.E. 2002. A theoretical study of the mechanism for peptide hydrolysis by thermolysin. JBIC Journal of Biological Inorganic Chemistry, 7, 284–298.CrossRefGoogle ScholarPubMed
Sharma, K.M., Kumar, R., Panwar, S. & Kumar, A. 2017. Microbial alkaline proteases: optimization of production parameters and their properties. Journal of Genetic Engineering and Biotechnology, 15, 115–126.CrossRefGoogle ScholarPubMed
Sizemore, R.K. & Stevenson, L.H. 1970. Method for the isolation of proteolytic marine bacteria. Applied Microbiology, 20, 991.CrossRefGoogle ScholarPubMed
Srimathi, S., Jayaraman, G. & Narayanan, P.R. 2006. Improved thermodynamic stability of subtilisin Carlsberg by covalent modification. Enzyme and Microbial Technology, 39, 301–307.CrossRefGoogle Scholar
Tallant, C., García-Castellanos, R., Seco, J., Baumann, U. & Gomis-Rüth, F.X. 2006. Molecular analysis of ulilysin, the structural prototype of a new family of metzincin metalloproteases. Journal of Biological Chemistry, 281, 17920–17928.CrossRefGoogle ScholarPubMed
Ward, O. 2011. Proteases. In Moo-Young, M., ed., Comprehensive biotechnology, 2nd edition. Cambridge, MA: Academic Press, 571–582.CrossRefGoogle Scholar