Bowman-birk proteinase inhibitor from Clitoria fairchildiarta seeds : isolation, biochemical properties and insecticidal potential
Mirian Dartzger, Ilka Maria Vasconcelos, valéria Scorsato, Ricardo Aparicio, Sergio Marangoni, Maria Lígia Rodrigues Macedo
ARTIGO
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Herein described is the biochemical characterisation, including in vitro and in vivo assays, for a proteinase inhibitor purified from Clitoria fairchildiana seeds (CFPI). Purification was performed by hydrophobic interaction and gel filtration chromatography. Kinetic studies of the purified...
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Herein described is the biochemical characterisation, including in vitro and in vivo assays, for a proteinase inhibitor purified from Clitoria fairchildiana seeds (CFPI). Purification was performed by hydrophobic interaction and gel filtration chromatography. Kinetic studies of the purified inhibitor showed a competitive-type inhibitory activity against bovine trypsin and chymotrypsin, with an inhibition stoichiometry of 1:1 for both enzymes. The inhibition constants against trypsin and chymotrypsin were 3.3 x 10(-10) and 1.5 x 10(-10) M, respectively, displaying a tight binding property. SDS-PAGE showed that CFPI has a single polypeptide chain with an apparent molecular mass of 15 kDa under non-reducing conditions. However, MALDI-TOF analysis demonstrated a molecular mass of 7.973 kDa, suggesting that CFPI is dimeric in solution. The N-terminal sequence of CFPI showed homology with members of the Bowman-Birk inhibitor family. CFPI remained stable to progressive heating for 30 min to each temperature range of 37 up to 100 degrees C and CD analysis exhibited no changes in spectra at 207 nm after heating at 90 degrees C and subsequent cooling. Moreover, CFPI was active over a wide pH range (2-10). In contrast, reduction with DTT resulted in a loss of inhibitory activity against trypsin and chymotrypsin. CFPI also exhibited significant inhibitory activity against larval midgut trypsin enzymes from Anagasta kuehniella (76%), Diatraea saccharalis (59%) and Heliothis virescens (49%). Its insecticidal properties were further analysed by bioassays and confirmed by negative impact on A. kuehniella development
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FUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPESP
2011/09361-0
CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO - CNPQ
142348/2010-4
FUNDAÇÃO DE APOIO AO DESENVOLVIMENTO DO ENSINO, CIÊNCIA E TECNOLOGIA DO ESTADO DE MATO GROSSO DO SUL - FUNDECT
FINANCIADORA DE ESTUDOS E PROJETOS - FINEP
fechado
Bowman-birk proteinase inhibitor from Clitoria fairchildiarta seeds : isolation, biochemical properties and insecticidal potential
Mirian Dartzger, Ilka Maria Vasconcelos, valéria Scorsato, Ricardo Aparicio, Sergio Marangoni, Maria Lígia Rodrigues Macedo
Bowman-birk proteinase inhibitor from Clitoria fairchildiarta seeds : isolation, biochemical properties and insecticidal potential
Mirian Dartzger, Ilka Maria Vasconcelos, valéria Scorsato, Ricardo Aparicio, Sergio Marangoni, Maria Lígia Rodrigues Macedo
Fontes
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Phytochemistry: the international journal of plant chemistry, plant biochemistry and molecular biology Vol. 118 (Oct., 2015), p. 224-235 |