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Purification and characterization of a trypsin inhibitor from Senna tora active against midgut protease of podborer

Paper ID Volume ID Publish Year Pages File Format Full-Text
34803 45048 2014 9 PDF Available
Title
Purification and characterization of a trypsin inhibitor from Senna tora active against midgut protease of podborer
Abstract

•This is the first report on characterization of Senna tora trypsin inhibitor (TI).•The inhibitor is monomeric with two isoforms [19,725 & ∼19,900 Da] having acidic pI.•The TI is stable in wide pH (2–12), temperature (30–100 °C) range with Ki 0.23 nM.•MALDI-PMF & N-Terminal sequence studies suggest it as previously unreported trypsin inhibitor.•Can be employed in pest management as it inhibited polyphagous pest gut proteases.

Proteinaceous protease inhibitors have potential application in medicines, agriculture and biotechnology. Present study was undertaken to purify and characterize a proteinaceous protease inhibitor from a medicinal plant, Senna tora syn. Cassia tora. The inhibitor was purified by ammonium sulphate precipitation, anion exchange (Q-sepharose), affinity (trypsin-sepharose) and molecular exclusion (sephadex G-75) chromatography. Zymography and denaturing polyacrylamide gel electrophoresis revealed a single band of ∼20 kDa trypsin inhibitor. Two dimensional polyacrylamide gel electrophoresis (2D-PAGE) and Matrix-assisted laser desorption ionization (MALDI) analyses revealed the presence of 19,725 Da (pI 4.60) and ∼19,900 Da (pI 4.57) isoform proteins in purified inhibitor. Protein identification by MALDI-peptide mass fingerprinting did not reveal high MASCOT (Matrix science) scores matching with previously known inhibitors. N-terminal amino acid sequence suggested this protein as a previously unreported inhibitor. Its dissociation constant (0.23 × 10−9 M) was indicative of a high affinity trypsin inhibitor. The inhibitor was stable over a broad range of pH (4–10) and temperature (30–60 °C). The purified inhibitor effectively inhibited total protease and trypsin-like activities of podborer (Helicoverpa armigera) midgut preparation. Hence, the inhibitor and its gene(s) can find application in combating against pest and protease dependent pathogens.

Keywords
Protease inhibitor; Helicoverpa armigera; Gut proteases; MALDI
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Purification and characterization of a trypsin inhibitor from Senna tora active against midgut protease of podborer
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Publisher
Database: Elsevier - ScienceDirect
Journal: Process Biochemistry - Volume 49, Issue 2, February 2014, Pages 347–355
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us