Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: DrugBank:EXPT01586 (G418)
 2,237 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Human renin is synthesized as a 406-amino acid preprorenin protein that is processed by a signal peptidase during secretion, to release prorenin as a 386-amino acid zymogen. The 46-amino acid "pro" domain is removed by a renin-processing enzyme, to produce enzymatically active renin, by cleavage at an Arg-Leu bond. The effects of the renin-processing enzyme can be mimicked by trypsin activation, where high concentrations of trypsin are incubated with prorenin for brief periods of time, followed by excess trypsin inhibitor to minimize secondary proteolytic processing by trypsin. In order to study the role of the pro segment in the secretion, folding, and activity of human renin, we engineered a construct where the pro domain from the preprorenin cDNA was deleted. This construct was introduced into mammalian cells and its expression was assayed in transient and stable systems. In COS-1 cells transfected with the prerenin expression vector pREN3, active renin was secreted with a specific activity of 1360 micrograms of angiotensin l/min/mg, compared with trypsin-activated prorenin, which has a specific activity of 818 micrograms of angiotensin l/min/mg. The active renin secreted in this system had a significantly reduced potency for the renin inhibitor SQ 32,970. These results demonstrate that the pro segment is dispensable for the folding and secretion of renin. A permanent cell line expressing the active form of renin was obtained by co-transfection of NRP cells with pREN3 and pHyg. A colony designated B/1 was identified, subcloned, and shown to secrete active renin (110 pg of renin/10(6) cells) optimally when maintained in both G418 and hygromycin.
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PMID:Stable expression, secretion, and characterization of active human renin in mammalian cells. 173 22

We have constructed a yeast integration vector targeted to chromosomal Ty delta sequences and used it to create Saccharomyces cerevisiae strains with stable tandem integrations ranging from 1 to 30 vector copies. The vector carries the bacterial NEO gene, allowing copy number to be tuned by varying G418 resistance, which generally increases with copy number as determined by quantitative Southern blot. Tandem integration into a single site is most commonly observed, but single-copy and two-site integration is also observed. Bovine pancreatic trypsin inhibitor was constitutively expressed and secreted using the NEO-based delta vector, and secretion levels were 2-10-fold improved relative to commonly used 2 mu multicopy yeast plasmids. The NEO-based Ty delta vector is a powerful tool for stable heterologous protein expression and secretion in yeast.
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PMID:An integrating vector for tunable, high copy, stable integration into the dispersed Ty delta sites of Saccharomyces cerevisiae. 884 5

The conversion of insulin-like growth factor-I (IGF-I) to the biologically more active des (1-3) IGF-I variant is catalyzed by a ubiquitous protease. This proteolytic activity is inhibited by human alpha1-antitrypsin and soy-bean trypsin inhibitor and is up-regulated in serum and tissue extracts of hypophysectomized rats. These observations lead us to investigate whether the growth hormone regulated, serine protease inhibitor, Spi 2.1 was able to inhibit the des (1-3) IGF-I generating protease. Dihydrofolate reductase deficient Chinese hamster ovary (CHO(dhfr-ve)) cells were transfected with a rat Spi 2.1 expression vector containing the dhfr and neomycin resistance gene. Stable transfectants were selected using G418 and amplified using methotrexate. Conditioned medium from Spi 2.1 transfected CHO cells potently inhibited proteolytic activity directed against a synthetic hexa-peptide with a sequence identical to the N-terminal of IGF-I. In contrast conditioned medium from wild-type CHO cells had little effect. Based upon these observations we suggest that our previous finding of enhanced des (1-3) IGF-I generating protease activity in growth hormone deficient rats may be, at least partly explained by reduced levels of Spi 2.1. Furthermore, we propose that the regulation of the generation of des (1-3) IGF-I may be an additional potential site of growth hormone regulation of IGF-I action.
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PMID:The growth hormone dependent serine protease inhibitor, Spi 2.1 inhibits the des (1-3) insulin-like growth factor-I generating protease. 938 51

Agrobacterium-mediated transformation was used to introduce a trypsin inhibitor gene into Taiwan cauliflower (Brassica oleracea var. botrytis L.) cultivars. The TI gene was isolated from a well-adapted Taiwan sweet potato cultivar and was expected to be especially effective in combating local pests. In vitro regeneration studies indicated that 4-day-old cauliflower seedling hypocotyl segments, pretreated with 2,4-dichlorophenoxyacetic acid for 3 days and incubated on a silver-ion-containing shoot induction medium, gave regeneration rates greater than 95%. Optimum transformation conditions were determined. G418 selection at 15 mg/l was initiated 1 week after cocultivation, and the dose was doubled 1 week later. Over 100 putative transgenic plants were produced. Transgenic status was confirmed by in vitro TI activity, and Southern and Western hybridization assays. The transgenic plants demonstrated in planta resistance to local insects to which the control plants were vulnerable.
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PMID:Development of insect-resistant transgenic cauliflower plants expressing the trypsin inhibitor gene isolated from local sweet potato. 3073 56