Record Details

Purification and characterization of Bowman-Birk and Kunitz isoinhibitors from the seeds of Rhynchosia sublobata (Schumach.) Meikle, a wild relative of pigeonpea

OAR@ICRISAT

View Archive Info
 
 
Field Value
 
Relation http://oar.icrisat.org/11087/
http://dx.doi.org/10.1016/j.phytochem.2018.12.018
10.1016/j.phytochem.2018.12.018
 
Title Purification and characterization of Bowman-Birk and Kunitz isoinhibitors from the seeds of Rhynchosia sublobata (Schumach.) Meikle, a wild relative of pigeonpea
 
Creator Mohanraj, S S
Gujjarlapudi, M
Lokya, V
Mallikarjuna, N
Dutta-Gupta, A
Padmasree, K
 
Subject Helicoverpa
Pigeonpea
Genetics and Genomics
 
Description Rhynchosia sublobata, a wild relative of pigeonpea, possesses defensive proteinase/protease inhibitors (PIs). Characterization of trypsin specific PIs (RsPI) separated from seeds by column chromatography using 2-D gel electrophoresis and Edman degradation method identified R. sublobata possessed both Bowman-Birk isoinhibitors (RsBBI) and Kunitz isoinhibitors (RsKI). A quick method was developed to separate RsBBI and RsKI from RsPI based on their differential solubility in TCA and acetate buffer. N-terminus sequencing of RsBBI and RsKI by MALDI-ISD ascertained the presence of Bowman Birk and Kunitz type isoinhibitors in R. sublobata. RsBBI (9216 Da) and RsKI (19,412 Da) exhibited self-association pattern as revealed by western blotting with anti-BBI antibody and MALDI-TOF peptide mass fingerprint analysis, respectively. RsBBI and RsKI varied significantly in their biochemical, biophysical and insecticidal properties. RsBBI inhibited the activity of trypsin (Ki = 128.5 ± 4.5 nM) and chymotrypsin (Ki = 807.8 ± 23.7 nM) while RsKI (Ki = 172.0 ± 9.2 nM) inhibited the activity of trypsin alone, by non-competitive mode. The trypsin inhibitor (TI) and chymotrypsin inhibitor (CI) activities of RsBBI were stable up to 100 °C. But, RsBBI completely lost its TI and CI activities on reduction with 3 mM DTT. Conversely, RsKI lost its TI activity on heating at 100 °C and retained >60% of its TI activity in presence of 3 mM DTT. CD spectroscopic studies on RsBBI and RsKI showed their secondary structural elements in the following order: random coils > β-sheets/β-turns > α-helix. However, RsKI showed reversible denaturation midpoint (Tm) of 75 °C. Further, the significant inhibitory activity of RsBBI (IC50 = 24 ng) and RsKI (IC50 = 59 ng) against trypsin-like gut proteases of Achaea janata (AjGPs) and Helicoverpa armigera (HaGPs) suggest them as potential biomolecules in the management of A. janata and H. armigera, respectively.
 
Publisher Elsevier
 
Date 2019
 
Type Article
PeerReviewed
 
Format application/pdf
 
Language en
 
Rights —
 
Identifier http://oar.icrisat.org/11087/1/Purification%20and%20characterization%20of%20Bowman-Birk%20and%20Kunitz%20isoinhibitors.pdf
Mohanraj, S S and Gujjarlapudi, M and Lokya, V and Mallikarjuna, N and Dutta-Gupta, A and Padmasree, K (2019) Purification and characterization of Bowman-Birk and Kunitz isoinhibitors from the seeds of Rhynchosia sublobata (Schumach.) Meikle, a wild relative of pigeonpea. Phytochemistry (TSI), 159. pp. 159-171. ISSN 00319422