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Enzymes Produced by Halotolerant Spore-Forming Gram-Positive Bacterial Strains Isolated From a Resting Habitat (Restinga de Jurubatiba) in Rio de Janeiro, Brazil: Focus on Proteases

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Abstract

The screening for hydrolases-producing, halotolerant, and spore-forming gram-positive bacteria from the root, rhizosphere, and non-rhizosphere soil of Blutaparon portulacoides, a plant found in the Restinga de Jurubatiba located at the northern region of Rio de Janeiro State, Brazil, resulted in the isolation of 22 strains. These strains were identified as Halobacillus blutaparonensis (n = 2), Oceanobacillus picturae (n = 5), and Oceanobacillus iheyensis (n = 15), and all showed the ability to produce different extracellular enzymes. A total of 20 isolates (90.9 %) showed activity for protease, 5 (22.7 %) for phytase, 3 (13.6 %) for cellulase, and 2 (9.1 %) for amylase. Some bacterial strains were capable of producing three (13.6 %) or two (9.1 %) distinct hydrolytic enzymes. However, no bacterial strain with ability to produce esterase and DNase was observed. The isolate designated M9, belonging to the species H. blutaparonensis, was the best producer of protease and also yielded amylase and phytase. This strain was chosen for further studies regarding its protease activity. The M9 strain produced similar amounts of protease when grown either without or with different NaCl concentrations (from 0.5 to 10 %). A simple inspection of the cell-free culture supernatant by gelatin-sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) revealed the presence of three major alkaline proteases of 40, 50, and 70 kDa, which were fully inhibited by phenylmethylsulfonyl fluoride (PMSF) and tosyl-l-phenylalanine chloromethyl ketone (TPCK) (two classical serine protease inhibitors). The secreted proteases were detected in a wide range of temperature (from 4 to 45 °C) and their hydrolytic activities were stimulated by NaCl (up to 10 %). The serine proteases produced by the M9 strain cleaved gelatin, casein, albumin, and hemoglobin, however, in different extensions. Collectively, these results suggest the potential use of the M9 strain in biotechnological and/or industrial processes.

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Acknowledgments

This study was supported by grants from the following Brazilian agencies: Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), Fundação de Amparo à Pesquisa no Estado do Rio de Janeiro (FAPERJ), and Fundação Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES). André L.S. Santos and Lucy Seldin were supported by CNPq and FAPERJ fellowships.

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Authors and Affiliations

  1. Departamento de Microbiologia Geral, Instituto de Microbiologia Paulo de Góes (IMPG), Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, Brazil

    Anderson Fragoso d. Santos, Clarissa Almeida Pacheco, Roberta d. Santos Valle, Lucy Seldin & André Luis Souza d. Santos

  2. Programa de Pós-Graduação em Bioquímica, Instituto de Química (IQ), Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, Brazil

    Anderson Fragoso d. Santos & André Luis Souza d. Santos

  3. Laboratório de Investigação de Peptidases (LIP), Departamento de Microbiologia Geral, Instituto de Microbiologia Paulo de Góes (IMPG), Bloco E-subsolo, sala 05, Centro de Ciências da Saúde (CCS), Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, RJ, 21941-902, Brazil

    André Luis Souza d. Santos

Authors
  1. Anderson Fragoso d. Santos
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  2. Clarissa Almeida Pacheco
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  3. Roberta d. Santos Valle
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  4. Lucy Seldin
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  5. André Luis Souza d. Santos
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Corresponding author

Correspondence to André Luis Souza d. Santos.

Additional information

Anderson Fragoso dos Santos and Clarissa Almeida Pacheco contributed equally to this work.

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d. Santos, A.F., Pacheco, C.A., Valle, R.d.S. et al. Enzymes Produced by Halotolerant Spore-Forming Gram-Positive Bacterial Strains Isolated From a Resting Habitat (Restinga de Jurubatiba) in Rio de Janeiro, Brazil: Focus on Proteases. Appl Biochem Biotechnol 174, 2748–2761 (2014). https://doi.org/10.1007/s12010-014-1223-5

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  • DOI: https://doi.org/10.1007/s12010-014-1223-5

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