@article { ISI:000341800200002,
	title = {Differential antibiosis against helicoverpa armigera exerted by distinct inhibitory repeat domains of capsicum annuum proteinase inhibitors},
	journal = {Phytochemistry},
	volume = {101},
	year = {2014},
	month = {MAY},
	pages = {16-22},
	publisher = {PERGAMON-ELSEVIER SCIENCE LTD},
	address = {THE BOULEVARD, LANGFORD LANE, KIDLINGTON, OXFORD OX5 1GB, ENGLAND},
	abstract = {<p>Plant defensive serine proteinase inhibitors (Pis) are known to have negative impact on digestive physiology of herbivore insects and thus have a crucial role in plant protection. Here, we have assessed the efficacy and specificity of three previously characterized inhibitory repeat domain (IRD) variants from Capsicum annuum PIs viz., IRD-7, -9 and -12 against gut proteinases from Helicoverpa armigera. Comparative study of in silico binding energy revealed that IRD-9 possesses higher affinity towards H. armigera serine proteinases as compared to IRD-7 and -12. H. armigera fed on artificial diet containing 5 TIU/g of recombinant IRD proteins exhibited differential effects on larval growth, survival rate and other nutritional parameters. Major digestive gut trypsin and chymotrypsin genes were down regulated in the IRD fed larvae, while few of them were up-regulated, this indicate alterations in insect digestive physiology. The results corroborated with proteinase activity assays and zymography. These findings suggest that the sequence variations among Pis reflect in their efficacy against proteinases in vitro and in vivo, which also could be used for developing tailor-made multi-domain inhibitor gene(s). (C) 2014 Elsevier Ltd. All rights reserved.</p>},
	keywords = {Capsicum annuum, Helicoverpa armigera, Inhibitory repeat domain, Proteases, Proteinase inhibitors},
	issn = {0031-9422},
	doi = {10.1016/j.phytochem.2014.01.011},
	author = {Joshi, Rakesh S. and Gupta, Vidya S. and Giri, Ashok P.}
}
@article {49189,
	title = {Engineering inhibitory repeat domains of Pin-II type proteinase inhibitors indicate their high structural-functional tolerance to mutagenesis},
	journal = {Biochemical and Biophysical Research Communications},
	volume = {735},
	year = {2024},
	month = {NOV },
	pages = {150808},
	type = {Article},
	abstract = {<p>
	Plant proteinase inhibitors (PIs) are critical in defending against biotic stress. Most PIs contain an inhibitory repeat domain (IRD), which serves as the functional component, displaying a high degree of sequence and structural conservation. In this study, we examined the structural and functional resilience of IRDs using a combination of computational modeling and experimental validation. We have taken an evolution-based approach to enhance the PIs effectiveness of two previously identified Capsicum annuum IRDs, IRD4 and IRD10. Through in silico site-saturation mutagenesis of IRD4 and IRD10, we identified key sites associated with enhanced PI activity for targeted mutagenesis. Binding energy predictions for a mutant IRD library, tested against target proteases, suggested that positions R11 and N32 in IRD4 and N32 and H33 in IRD10 were promising candidates for further modification to improve inhibitory potential. Subsequent experimental validation revealed that the mutant proteins IRD4_R11K and IRD4_N32S exhibited stronger chymotrypsin inhibition than the wild-type (WT) IRD4. Similarly, the mutants IRD10_N32S and IRD10_H33 N demonstrated improved trypsin inhibition relative to the WT IRD10. These findings indicate that engineered IRD variants can tolerate structural changes while maintaining or enhancing their inhibitory activity against target proteases. Overall, this study demonstrates the potential of engineering PIs to increase their structural and functional resilience, offering new opportunities for biotechnological applications.</p>
},
	keywords = {Capsicum annum, Inhibitory repeat domain, molecular modeling, plant proteinase inhibitors, Proteases, Saturation mutagenesis},
	issn = {0006-291X},
	doi = {10.1016/j.bbrc.2024.150808},
	author = {Barbole, Ranjit S. and Joshi, Rakesh S. and Giri, Ashok P.}
}
