African Journal of
Biotechnology

  • Abbreviation: Afr. J. Biotechnol.
  • Language: English
  • ISSN: 1684-5315
  • DOI: 10.5897/AJB
  • Start Year: 2002
  • Published Articles: 12236

Full Length Research Paper

Kinetic characterization and molecular modeling of trehalose-6-phosphate phosphatase from Anopheles gambiae and expressed in Pichia pastoris

Marcos Cézar Fernandes Pessoa
  • Marcos Cézar Fernandes Pessoa
  • Laboratory of DNA Technology, Biotechnology Department, Multidisciplinary Support Center, Federal University of Amazonas, Manaus, Amazonas, Brazil.
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Érika Izumi
  • Érika Izumi
  • Federal University of Technology, Curitiba, Paraná, Brazil.
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Sandra Patrícia Zanotto
  • Sandra Patrícia Zanotto
  • Laboratory of DNA Technology, Biotechnology Department, Multidisciplinary Support Center, Federal University of Amazonas, Manaus, Amazonas, Brazil.
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Spartaco Astolfi-Filho
  • Spartaco Astolfi-Filho
  • Laboratory of DNA Technology, Biotechnology Department, Multidisciplinary Support Center, Federal University of Amazonas, Manaus, Amazonas, Brazil.
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  •  Received: 04 April 2017
  •  Accepted: 14 June 2017
  •  Published: 21 June 2017

Abstract

Trehalose-6-phosphate phosphatase (TPP) is one of the primary enzymes involved in the synthesis of trehalose, the main sugar found in insect hemolymph. In the present study, we report for the first time heterologous expression in Pichia pastoris, characterization and homology modeling of TPP from Anopheles gambiae mosquito. Purified TPP recombinant exhibited a molecular weight of approximately 36 kDa with optimum pH of 8.0, optimum temperature of 38°C, and the KM was 3.19 ± 0.10 mM. Inhibition tests revealed that CaCl2 and Ca(NO3)2 at 5 and 25 mM, respectively, were effective inhibitors of TPP activity. Homology studies and molecular modeling indicated high similarity of the TPP tridimensional structure with TPP enzymes deposited at a data bank and these homology studies confirmed that it is a trehalose phosphatase with conserved motifs of the superfamily haloacid dehydrogenase. These results may provide support for the discovering of TPP inhibitors to be used for development of insecticides to control mosquito vectors.

 

Key words: Trehalose, trehalose-6-phosphate phosphatase, expression, enzyme characterization, homology modeling.