Expression of the Klebsiella pneumoniae pullulanase-encoding gene in Saccharomyces cerevisiae
Date
1993
Authors
Janse B.J.H.
Pretorius I.S.
Journal Title
Journal ISSN
Volume Title
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Abstract
A 3800-base pair (bp) DNA fragment encoding the mature pullulanase from Klebsiella pneumoniae was inserted between two different yeast expression-secretion cassettes and an yeast gene terminator. These cassettes were cloned into an yeast centromeric plasmid YCplacIII and transformed into laboratory strains of Saccharomyces cerevisiae. Transcription initiation signals were derived from the mating pheromone α-factor (MFα1(p)) and alcohol dehydrogenase (ADC1(p)) gene promoters. Secretion of pullulanase was directed by the leader sequence of the yeast mating pheromone α-factor (MFα1(s)). Transcription termination was effected by the yeast tryptophan synthase gene terminator (TRP5(r)). Southernblot analysis confirmed the presence of pulA in transformed yeasts and Northern-blot analysis revealed the presence of PUL1 mRNA. A pullulan agarose assay indicated the extracellular production of biologically active pullulanase by S. cerevisiae.
Description
Keywords
messenger rna, pullulanase, article, cloning vector, enzyme assay, enzyme release, gene expression, gene insertion, klebsiella pneumoniae, molecular cloning, nonhuman, northern blotting, plasmid, priority journal, promoter region, saccharomyces cerevisiae, southern blotting, transcription initiation, transcription termination, Base Sequence, Blotting, Northern, Blotting, Southern, Cloning, Molecular, DNA, Fungal, Electrophoresis, Polyacrylamide Gel, Genes, Fungal, Glycoside Hydrolases, Klebsiella pneumoniae, Molecular Sequence Data, Peptides, Polymerase Chain Reaction, Restriction Mapping, Saccharomyces cerevisiae, Terminator Regions (Genetics), Transcription, Genetic, Transformation, Genetic, Tryptophan Synthase
Citation
Current Genetics
24
02-Jan
24
02-Jan