Title
A trypsin inhibitor purified from Cassia leiandra seeds has insecticidal activity against Aedes aegypti
Author
Lucas P. Dias, Jose T.A. Oliveira, Lady C.B. Rocha-Bezerra, Daniele O.B. Sousa, Helen P.S. Costa, Nadine M.S. Araujo, Ana F.U. Carvalho, Pedro M.S. Tabosa, Ana C.O. Monteiro-Moreira, Marina D.P. Lobo, Frederico B.M.B. Moreno, Bruno A.M. Rocha, José L.S. Lopes, Leila M. Beltramini, Ilka M. Vasconcelos
Year
2017
Journal
Process Biochemistry
Abstract
A trypsin inhibitor from C. leiandra seeds, named ClTI, was purified, characterized, and its insecticidal activity against Ae. aegypti evaluated. ClTI was purified by DEAE-Cellulose and trypsin-Sepharose 4B chromatography, with a 15.5-fold purification and 2.4% yield. ClTI is composed of a 19,484 Da polypeptide chain as revealed by mass spectrometry, it is not a glycoprotein, its amino acid sequence is similar to other Kunitz-type inhibitors, and it comprises 35% β-sheets, 14% β-turns, and 50% disordered secondary structures. ClTI is an uncompetitive inhibitor of bovine trypsin (IC50 of 33.81 × 10−8 M, Ki of 6.25 × 10−8 M) stable over a broad range of pHs (2.2–10.0) and temperatures (30–70 °C), but dithiothreitol led to a partial loss of the inhibitory activity. ClTI, at 4.65 × 10−6 M, reduced in 50% the activity of the Ae. aegypti midgut proteases. ClTI also promoted acute toxicity on the 3rd instar larvae of Ae. aegypti, with an LC50 of 2.28 × 10−2 M. Moreover, it caused a 24-h delay of the larvae development and 44% mortality after ten days of exposure. Altogether, these results suggest that ClTI has potential as a natural compound to control Ae. aegypti, a vector of several infection diseases.
Instrument
J-715
Keywords
Circular dichroism, Secondary structure, Thermal stability, Chemical stability, Biochemistry