Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.2.1.23 (beta-galactosidase)
 14,648 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Strategies for the expression of precursors of eukaryotic secreted proteins as part of fused proteins in Escherichia coli have been explored. A fusion protein with beta-galactosidase at the N-terminal end and honeybee prepromelittin at the C-terminal end (beta-gal-pM) was expressed in low amounts as a cleaved polypeptide, from which the promelittin portion had been removed. Inclusion in the induction culture of 10 mM MgCl2 or 8.3% (v/v) ethanol, inhibitors of signal peptidase, gave rise to the full-length beta-gal-pM fusion protein. The results suggest that a soluble recombinant fusion protein with a signal peptide in an internal location 660 residues from the N-terminus is recognized by the E. coli translocation apparatus in the inner membrane and by leader peptidase. High-level production (about 45% of total cellular proteins) of prepromelittin was achieved when it was part of a fusion protein at the C-terminus of a truncated insoluble polypeptide from bacteriophage gene 10. This fusion protein separated into inclusion bodies in an aggregated form. In contrast, attempts to express prepromelittin by itself or at the N-terminal end of a fusion with mouse dihydrofolate reductase (pM-DHFR) proved unsuccessful.
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PMID:Expression of honeybee prepromelittin as a fusion protein in Escherichia coli. 182 10

We have shown that the Leishmania major transfection vector pR-NEO (or derivatives thereof) can be introduced and stably maintained in four species complexes of pathogenic Leishmania (L. tropica, L. mexicana, L. donovani, L. braziliensis), and the genera Endotrypanum and Crithidia; transfection of Trypanosoma cruzi or Trypanosoma brucei was not successful. Quantitative plating assays showed that the transfection efficiencies were high in L. major and Leishmania amazonensis (5x10(-5)/cell) and about 10-fold less for Leishmania panamaensis and Crithidia. Leishmania donovani transfected with pR-NEO retained the ability to infect hamsters, and amastigotes recovered after 2 months yielded G418-resistant promastigotes which retained high levels of extrachromosomal pR-NEO DNA. In promastigotes, the transfected DNA existed as extrachromosomal circles, and expressed the predicted 2.4-kb hybrid NEO/DHFR-TS mRNA bearing the trans-spliced miniexon. Large quantitative differences were observed only in Crithidia: relative to transfected Leishmania species, the copy number of pR-NEO was elevated 20-fold, while the levels of the NEO/DHRFR-TS mRNA or Escherichia coli beta-galactosidase (synthesized from the expression vector pX-beta GAL) were reduced 80 and more than 1000-fold, respectively. Thus, genetic signals derived from L. major DNA that mediate RNA expression or stability are recognized by the heterologous Leishmania species but less efficiently by Crithidia. These studies suggest that pR-NEO derived vectors may be applied to the study of genes expressed throughout the life cycle in a wide range of pathogenic trypanosomatids.
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PMID:Stable DNA transfection of a wide range of trypanosomatids. 190 80

Type III machines of pathogenic Yersinia spp. transport Yop proteins across the bacterial envelope into host cells. Translational fusions of yopE to the dihydrofolate reductase gene (dhfr) or the beta-galactosidase gene (lacZ) generate hybrid proteins that block type III injection of Yop proteins into host cells, consistent with the canonical view that impassable DHFR and LacZ hybrids jam secretion machines. Mutations in repressors of posttranscriptional gene regulation, Yersinia enterocolitica yscM1 and yscM2 as well as Yersinia pestis lcrQ, relieve the YopE-DHFR-imposed blockade and restore type III injection into host cells. Genetic suppression of the type III blockade does not, however, promote YopE-DHFR secretion. A model is proposed whereby rejection of YopE-DHFR from the secretion pathway inhibits type III gene expression.
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PMID:Rejection of impassable substrates by Yersinia type III secretion machines. 1619 80

The rat parasite Eimeria nieschulzi is a suitable model for transfection studies and was used as an additional model organism for the genus Eimeria. We describe the transfection of this apicomplexan parasites and the cultivation of transformed stages in cell culture and in vivo. The beta-galactosidase or yellow fluorescent protein was expressed in all parasitic stages up to the second merozoite generation in vitro under control of the heterologous promoter region of Eimeria tenella mic1 gene previously described for E. tenella transfection. Pyrimethamine resistant E. nieschulzi parasites were obtained in vitro after transfection with a plasmid encoding the Toxoplasma gondii dhfr/ts-m2m3 gene. Co-transfection experiments with an YFP-plasmid resulted in pyrimethamine resistant and fluorescent parasitic stages. Infection of rats with transfected E. nieschulzi sporozoites directed to expression of beta-galactosidase or YFP in oocysts. Co-transfection with YFP/DHFR-TS allowed selection of resistant parasites in vivo. Excreted transgenic oocysts showed arrangement of YFP expression which lead to questions about meiotic recombination frequency and mechanisms.
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PMID:Reporter gene expression in cell culture stages and oocysts of Eimeria nieschulzi (Coccidia, Apicomplexa). 1879 26