Dataset Information

Evaluating the role of a trypsin inhibitor from soap nut (Sapindus trifoliatus L. Var. Emarginatus) seeds against larval gut proteases, its purification and characterization.

ABSTRACT: The defensive capacities of plant protease Inhibitors (PI) rely on inhibition of proteases in insect guts or those secreted by microorganisms; and also prevent uncontrolled proteolysis and offer protection against proteolytic enzymes of pathogens.An array of chromatographic techniques were employed for purification, homogeneity was assessed by electrophoresis. Specificity, Ki value, nature of inhibition, complex formation was carried out by standard protocols. Action of SNTI on insect gut proteases was computationally evaluated by modeling the proteins by threading and docking studies by piper using Schrodinger tools.We have isolated and purified Soap Nut Trypsin Inhibitor (SNTI) by acetone fractionation, ammonium sulphate precipitation, ion exchange and gel permeation chromatography. The purified inhibitor was homogeneous by both gel filtration and polyacrylamide gel electrophoresis (PAGE). SNTI exhibited a molecular weight of 29 kDa on SDS-PAGE, gel filtration and was negative to Periodic Acid Schiff's stain. SNTI inhibited trypsin and pronase of serine class. SNTI demonstrated non-competitive inhibition with a Ki value of 0.75?±?0.05×10-10 M. The monoheaded inhibitor formed a stable complex in 1:1 molar ratio. Action of SNTI was computationally evaluated on larval gut proteases from Helicoverpa armigera and Spodoptera frugiperda. SNTI and larval gut proteases were modeled and docked using Schrodinger software. Docking studies revealed strong hydrogen bond interactions between Lys10 and Pro71, Lys299 and Met80 and Van Der Waals interactions between Leu11 and Cys76amino acid residues of SNTI and protease from H. Armigera. Strong hydrogen bonds were observed between SNTI and protease of S. frugiperda at positions Thr79 and Arg80, Asp90 and Gly73, Asp2 and Gly160 respectively.We conclude that SNTI potentially inhibits larval gut proteases of insects and the kinetics exhibited by the protease inhibitor further substantiates its efficacy against serine proteases.

SUBMITTER: Gandreddi VD

PROVIDER: S-EPMC4618930 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Evaluating the role of a trypsin inhibitor from soap nut (Sapindus trifoliatus L. Var. Emarginatus) seeds against larval gut proteases, its purification and characterization.

Gandreddi V D Sirisha VD Kappala Vijaya Rachel VR Zaveri Kunal K Patnala Kiranmayi K

BMC biochemistry 20151022

<h4>Background</h4>The defensive capacities of plant protease Inhibitors (PI) rely on inhibition of proteases in insect guts or those secreted by microorganisms; and also prevent uncontrolled proteolysis and offer protection against proteolytic enzymes of pathogens.<h4>Methods</h4>An array of chromatographic techniques were employed for purification, homogeneity was assessed by electrophoresis. Specificity, Ki value, nature of inhibition, complex formation was carried out by standard protocols. ...[more]

PMID: 26489418

Similar Datasets

Project description:BackgroundBlood flukes (Schistosoma spp.) are parasites that can survive for years or decades in the vasculature of permissive mammalian hosts, including humans. Proteolytic enzymes (proteases) are crucial for successful parasitism, including aspects of invasion, maturation and reproduction. Most attention has focused on the 'cercarial elastase' serine proteases that facilitate skin invasion by infective schistosome larvae, and the cysteine and aspartic proteases that worms use to digest the blood meal. Apart from the cercarial elastases, information regarding other S. mansoni serine proteases (SmSPs) is limited. To address this, we investigated SmSPs using genomic, transcriptomic, phylogenetic and functional proteomic approaches.Methodology/principal findingsGenes encoding five distinct SmSPs, termed SmSP1 - SmSP5, some of which comprise disparate protein domains, were retrieved from the S. mansoni genome database and annotated. Reverse transcription quantitative PCR (RT- qPCR) in various schistosome developmental stages indicated complex expression patterns for SmSPs, including their constituent protein domains. SmSP2 stood apart as being massively expressed in schistosomula and adult stages. Phylogenetic analysis segregated SmSPs into diverse clusters of family S1 proteases. SmSP1 to SmSP4 are trypsin-like proteases, whereas SmSP5 is chymotrypsin-like. In agreement, trypsin-like activities were shown to predominate in eggs, schistosomula and adults using peptidyl fluorogenic substrates. SmSP5 is particularly novel in the phylogenetics of family S1 schistosome proteases, as it is part of a cluster of sequences that fill a gap between the highly divergent cercarial elastases and other family S1 proteases.Conclusions/significanceOur series of post-genomics analyses clarifies the complexity of schistosome family S1 serine proteases and highlights their interrelationships, including the cercarial elastases and, not least, the identification of a 'missing-link' protease cluster, represented by SmSP5. A framework is now in place to guide the characterization of individual proteases, their stage-specific expression and their contributions to parasitism, in particular, their possible modulation of host physiology.

Dataset Information

Evaluating the role of a trypsin inhibitor from soap nut (Sapindus trifoliatus L. Var. Emarginatus) seeds against larval gut proteases, its purification and characterization.

Publications

Evaluating the role of a trypsin inhibitor from soap nut (Sapindus trifoliatus L. Var. Emarginatus) seeds against larval gut proteases, its purification and characterization.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets