title: Inhibitory effects of a Kunitz-type inhibitor from Pithecellobium dumosum (Benth) seeds against insect-pests' digestive proteinases.
abstract: Pithecellobium dumosum is a tree belonging to the Mimosoideae subfamily that presents various previously characterized Kunitz-type inhibitors. The present study provides a novel Kunitz-trypsin inhibitor isoform purified from P. dumosum seeds. Purification procedure was performed by TCA precipitation followed by a trypsin-Sepharose chromatography and a further reversed-phase HPLC. Purified inhibitor (PdKI-4) showed enhanced inhibitory activity against bovine trypsin and chymotrypsin. Furthermore, PdKI-4 showed remarkable inhibitory activity against serine proteases from the coleopterans Callosobruchus maculatus and Zabrotes subfasciatus, and the lepidopterans Alabama argillacea and Telchin licus. However, PdKI-4 was unable to inhibit porcine pancreatic elastase, pineapple bromelain and Carica papaya papain. SDS-PAGE showed that PdKI-4 consisted of a single polypeptide chain with molecular mass of 21 kDa. Kinetic studies demonstrated that PdKI-4 is probably a competitive inhibitor with a Ki value of 5.7 × 10(-10) M for bovine trypsin. PdKI-4 also showed higher stability over a wide range of temperature (37-100 °C) and pH (2-12). N-termini sequence was obtained by Edman degradation showing higher identity with other Mimosoideae subfamily Kunitz-type inhibitor members. In summary, data here reported indicate the biotechnological potential of PdKI-4 for development of products against insect-pests.
authors.affiliation: Depto de Bioquímica, CB, UFRN, Natal, RN, Brazil.
authors.forename: Fabiola P S
authors.initials: FP
authors.lastname: Rufino
authors.forename: Vanessa M A
authors.initials: VM
authors.lastname: Pedroso
authors.forename: Jonalson N
authors.initials: JN
authors.lastname: Araujo
authors.forename: Anderson F J
authors.initials: AF
authors.lastname: França
authors.forename: Luciana M A
authors.initials: LM
authors.lastname: Rabêlo
authors.forename: Ludovico
authors.initials: L
authors.lastname: Migliolo
authors.forename: Sumika
authors.initials: S
authors.lastname: Kiyota
authors.forename: Elizeu A
authors.initials: EA
authors.lastname: Santos
authors.forename: Octavio L
authors.initials: OL
authors.lastname: Franco
authors.forename: Adeliana S
authors.initials: AS
authors.lastname: Oliveira
mesh_terms: D000818:Animals; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D007887:Fabaceae / Q000737:chemistry*; D007313:Insecta / Q000201:enzymology*; D010447:Peptide Hydrolases / Q000378:metabolism*; D010455:Peptides / Q000494:pharmacology*; D010940:Plant Proteins / Q000494:pharmacology*; D012639:Seeds / Q000737:chemistry*
chemical_list: D004791:Enzyme Inhibitors; C053865:Kunitz-type protease inhibitor, plant; D010455:Peptides; D010940:Plant Proteins; D010447:Peptide Hydrolases