title: Purification of a 6.5 kDa protease inhibitor from Amazon Inga umbratica seeds effective against serine proteases of the boll weevil Anthonomus grandis.
abstract: A 6.5 kDa serine protease inhibitor was purified by anion-exchange chromatography from the crude extract of the Inga umbratica seeds, containing inhibitor isoforms ranging from 6.3 to 6.7 kDa and protease inhibitors of approximately 19 kDa. The purified protein was characterized as a potent inhibitor against trypsin and chymotrypsin and it was named I. umbratica trypsin and chymotrypsin inhibitor (IUTCI). MALDI-TOF spectra of the IUTCI, in the presence of DTT, showed six disulfide bonds content, suggesting that this inhibitor belongs to Bowman-Birk family. The circular dichroism spectroscopy indicates that IUTCI is predominantly formed by unordered and beta-sheet secondary structure. It was also characterized, by fluorescence spectroscopy, as a stable protein at range of pH from 5.0 to 7.0. Moreover, this inhibitor at concentration of 75 microM presented a remarkable inhibitory activity (60%) against digestive serine proteases from boll weevil Anthonomus grandis, an important economical cotton pest.
authors.affiliation: Universidade de Brasília, Departamento de Biologia Celular, Brasília, DF, Brazil. nina@unb.br.
authors.forename: L A
authors.initials: LA
authors.lastname: Calderon
authors.forename: R C L
authors.initials: RC
authors.lastname: Teles
authors.forename: J R S A
authors.initials: JR
authors.lastname: Leite
authors.forename: O L
authors.initials: OL
authors.lastname: Franco
authors.forename: M F
authors.initials: MF
authors.lastname: Grossi-de-Sá
authors.forename: F J
authors.initials: FJ
authors.lastname: Medrano
authors.forename: C
authors.initials: C
authors.lastname: Bloch
authors.forename: S M
authors.initials: SM
authors.lastname: Freitas
mesh_terms: D002852:Chromatography, Ion Exchange; D002942:Circular Dichroism; D007887:Fabaceae / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D008970:Molecular Weight; D012639:Seeds / Q000737:chemistry*; D012697:Serine Endopeptidases / Q000378:metabolism*; D015842:Serine Proteinase Inhibitors / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology*; D013050:Spectrometry, Fluorescence; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D048889:Weevils / Q000201:enzymology*
chemical_list: D015842:Serine Proteinase Inhibitors; D012697:Serine Endopeptidases