| Full text | |
| Author(s): |
Dantzger, Miriam
[1, 2]
;
Vasconcelos, Ilka Maria
[3]
;
Scorsato, Valeria
[4, 5]
;
Aparicio, Ricardo
[4]
;
Marangoni, Sergio
[1]
;
Rodrigues Macedo, Maria Ligia
[1, 2]
Total Authors: 6
|
| Affiliation: | [1] Univ Estadual Campinas, Inst Biol, Dept Biochem, BR-13083970 Campinas, SP - Brazil
[2] Univ Mato Grosso Sul, Ctr Biol & Hlth Sci, Dept Food Technol & Publ Hlth, BR-79070900 Campo Grande, MS - Brazil
[3] Univ Ceara, Dept Biochem & Mol Biol, BR-60451970 Fortaleza, CE - Brazil
[4] Univ Estadual Campinas, Inst Chem, Lab Struct Biol & Crystallog, BR-13083970 Campinas, SP - Brazil
[5] Univ Estadual Campinas, Inst Biol, BR-13083970 Campinas, SP - Brazil
Total Affiliations: 5
|
| Document type: | Journal article |
| Source: | Phytochemistry; v. 118, p. 224-235, OCT 2015. |
| Web of Science Citations: | 9 |
| Abstract | |
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. (C) 2015 Elsevier Ltd. All rights reserved. (AU) | |
| FAPESP's process: | 11/09361-0 - Entada acaciifolia trypsin inhibitor: structural characterization and evaluation of insecticide potential against the fall armyworm (Spodoptera frugiperda) |
| Grantee: | Sergio Marangoni |
| Support type: | Regular Research Grants |