Cloning and High-Level Expression of α-Galactosidase cDNA fromPenicillium purpurogenum
| Autor: | Isao Kusakabe, Gwi Gun Park, Satoshi Kaneko, Hajime Shibuya, Shigeki Yoshida, Hiroaki Nagasaki, Hideyuki Kobayashi |
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| Rok vydání: | 1998 |
| Předmět: |
DNA
Complementary Molecular Sequence Data Penicillium purpurogenum Oligosaccharides Sequence alignment Molecular cloning Applied Microbiology and Biotechnology Complementary DNA Carbohydrate Conformation Humans Amino Acid Sequence Cloning Molecular Enzymology and Protein Engineering Peptide sequence Trichoderma reesei DNA Primers chemistry.chemical_classification Base Sequence Sequence Homology Amino Acid Ecology biology Reverse Transcriptase Polymerase Chain Reaction Penicillium Nucleic acid sequence biology.organism_classification Molecular biology Recombinant Proteins Amino acid Carbohydrate Sequence chemistry Biochemistry alpha-Galactosidase Sequence Alignment Food Science Biotechnology |
| Zdroj: | Applied and Environmental Microbiology. 64:4489-4494 |
| ISSN: | 1098-5336 0099-2240 |
| Popis: | The cDNA coding forPenicillium purpurogenumα-galactosidase (αGal) was cloned and sequenced. The deduced amino acid sequence of the α-Gal cDNA showed that the mature enzyme consisted of 419 amino acid residues with a molecular mass of 46,334 Da. The derived amino acid sequence of the enzyme showed similarity to eukaryotic αGals from plants, animals, yeasts, and filamentous fungi. The highest similarity observed (57% identity) was toTrichoderma reeseiAGLI. The cDNA was expressed inSaccharomyces cerevisiaeunder the control of the yeastGAL10promoter. Almost all of the enzyme produced was secreted into the culture medium, and the expression level reached was approximately 0.2 g/liter. The recombinant enzyme purified to homogeneity was highly glycosylated, showed slightly higher specific activity, and exhibited properties almost identical to those of the native enzyme fromP. purpurogenumin terms of the N-terminal amino acid sequence, thermoactivity, pH profile, and mode of action on galacto-oligosaccharides. |
| Databáze: | OpenAIRE |
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