@article { ISI:000250610300004,
	title = {Diverse forms of pin-II family proteinase inhibitors from capsicum annuum adversely affect the growth and development of helicoverpa armigera},
	journal = {Gene},
	volume = {403},
	number = {1-2},
	year = {2007},
	month = {NOV},
	pages = {29-38},
	publisher = {ELSEVIER SCIENCE BV},
	type = {Article},
	address = {PO BOX 211, 1000 AE AMSTERDAM, NETHERLANDS},
	abstract = {<p>Novel forms of Pin-II type proteinase inhibitor (PIs) cDNAs (CanPIs) having three or four inhibitory repeat domains (IRD) were isolated from the developing green fruits of Capsicum annuum. Deduced amino acid (aa) sequences of the CanPIs showed up to 15\% sequence divergence among each other or reported inhibitors (CanPI-1 AF039398, CanPI-2 AF221097). Amino acid sequence analysis of these CanPIs revealed that three IRD Pls have trypsin inhibitory sites, while four IRD CanPIs have both trypsin and chymotrypsin inhibitory sites. Four CanPIs, two having three IRD (CanPI-3 AY986465 and CanPI-5 DQ005912) and two having four IRD (CanPI-7 DQ005913 and CanPI-9 DQ005915), were cloned in Pichia pastoris to express recombinant CanPIs. Recombinant CanPIs inhibited 90\% of bovine trypsin (TI), while chymotrypsin inhibition (0) varied with the number of chymotrypsin inhibitory sites in the CanPIs. Recombinant inhibitors inhibited over 70\% of the gut proteinase activity of Helicoverpa armigera. H. armigera larvae fed on recombinant CanPIs individually incorporated into artificial diet, showed 35\% mortality; in addition, weight gain in H. armigera larvae and pupae was severely reduced compared to controls. Of the four CanPIs, CanPI-7, which has two sites for TI and CI, was the only one to have a consistently antagonistic effect on H. armigera growth and development. We conclude that among the four recombinant Pls tested, CanPIs containing diverse IRDs are best suited for developing insect-resistant transgenic plants. (c) 2007 Elsevier B.V. All rights reserved.</p>},
	keywords = {insect resistance, IP repeats, PI-insect interaction, Pichia pastoris, potato type 2 inhibitors (Pot-II), Solanaceae},
	issn = {0378-1119},
	doi = {10.1016/j.gene.2007.07.024},
	author = {Tamhane, Vaijayantl A. and Giri, Ashok P. and Sainani, Mohini N. and Gupta, Vidya S.}
}
@article {45484,
	title = {Zebrafish acid ceramidase: expression in Pichia pastoris GS115and biochemical characterization},
	journal = {International Journal of Biological Macromolecules},
	volume = {122},
	year = {2019},
	month = {FEB},
	pages = {587-593},
	type = {Article},
	abstract = {<p>Acid ceramidase (N-acylsphingosine deacylase EC 3.5.1.23; AC) catalyzes the hydrolysis of ceramide into sphingosine (SPH) and free fatty acid. Zebrafish acid ceramidase (AC) has 60\% homology with the human AC). Mutations in the human AC gene asah1 are known to cause Farber disease and spinal muscular atrophy with progressive myoclonic epilepsy. Zebrafish AC was overexpressed in Pichia pastoris by inserting asah1b gene into the genome. The majority of the overexpressed enzyme was secreted into the culture medium and purified to apparent homogeneity by stepwise chromatography. The recombinant protein was glycosylated precursor, that further undergoes limited autoproteolytic processing into two subunits (alpha and beta) which are visible in SDS-PAGE. The zebrafish AC is heterodimer associated with an inter-subunit disulfide bond. SDS-PAGE estimated the mass of native enzyme to be approximately 50 kDa \&amp; size exclusion chromatography estimated the mass of the active enzyme as approximately 100 kDa, suggesting the formation of a dimer of heterodimers. The protein was secreted as a mixture of processed and unprocessed forms in the culture media. A preliminary characterization of purified zebrafish AC was done by an enzyme assay. The zebrafish AC expressed in Pichia pastoris would be used for further structural and functional analysis. (C) 2018 Elsevier B.V. All rights reserved.</p>},
	keywords = {Autoproteolytic processing, Pichia pastoris, Zebrafish acid ceramidase},
	issn = {0141-8130},
	doi = {10.1016/j.ijbiomac.2018.10.222},
	author = {Rajput, Vijay B. and Karthikeyan, Muthukumarasamy and Ramasamy, Sureshkumar}
}
