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  • 1995-1999  (4)
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  • 1
    Digitale Medien
    Digitale Medien
    Cloning, sequence and expression of the gene coding for rhamnogalacturonase of Aspergillus aculeatus; a novel pectinolytic enzyme (1995)
    Springer
    Applied microbiology and biotechnology 43 (1995), S. 861-870 
    Details
    ISSN: 1432-0614
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie , Werkstoffwissenschaften, Fertigungsverfahren, Fertigung
    Notizen: Abstract  Rhamnogalacturonase was purified from culture filtrate of Aspergillus aculeatus after growth in medium with sugar-beet pulp as carbon source. Purified protein was used to raise antibodies in mice and with the antiserum obtained a gene coding for rhamnogalacturonase (rhgA) was isolated from a λ cDNA expression library. The cloned rhgA gene has an open-reading frame of 1320 base pairs encoding a protein of 440 amino acids with a predicted molecular mass of 45 962 Da. The protein contains a potential signal peptidase cleavage site behind Gly-18 and three potential sites for N-glycosylation. Limited homology with A. niger polygalacturonase amino acid sequences is found. A genomic clone of rhgA was isolated from a recombinant phage λ genomic library. Comparison of the genomic and cDNA sequences revealed that the coding region of the gene is interrupted by three introns. Furthermore, amino acid sequences of four different peptides, derived from purified A. aculeatus rhamnogalacturonase, were also found in the deduced amino acid sequence of rhgA. A. aculeatus strains overexpressing rhamnogalacturonase were obtained by cotransformation using either the A. niger pyrA gene or the A. aculeatus pyrA gene as selection marker. For expression of rhamnogalacturonase in A. awamori the A. awamori pyrA gene was used as selection marker. Degradation patterns of modified hairy regions, determined by HPLC, show the recombinant rhamnogalacturonase to be active, and the enzyme was found to have a positive effect in the apple hot-mash liquefaction process.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/s002530050497
    Bibliothek Standort Signatur Band/Heft/Jahr Verfügbarkeit
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  • 2
    Digitale Medien
    Digitale Medien
    An expression system based on the promoter region of the Aspergillus awamori 1,4-β-endoxylanase A gene (1996)
    Springer
    Applied microbiology and biotechnology 46 (1996), S. 28-35 
    Details
    ISSN: 1432-0614
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie , Werkstoffwissenschaften, Fertigungsverfahren, Fertigung
    Notizen: Abstract  A new, highly inducible fungal promoter derived from the Aspergillus awamori 1,4-β-endoxylanase A (exlA) gene is described. Induction analysis, carried out with the wild-type strain in shake flasks, showed that exlA expression is regulated at the transcriptional level. Using a β-glucuronidase (uidA) reporter strategy, D-xylose was shown to be an efficient inducer of the exlA promoter, whereas sucrose or maltodextrin were not. Upon D-xylose induction, the exlA promoter was threefold more efficient than the frequently used A. niger glucoamylase (glaA) promoter under maltodextrin induction. Detailed induction analyses demonstrated that induction was dependent on the presence of D-xylose in the medium. Carbon-source-limited chemostat cultures with the uidA reporter strain showed that D-xylose was also a very good inducer in a fermenter, even in the presence of sucrose.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/s002530050779
    Bibliothek Standort Signatur Band/Heft/Jahr Verfügbarkeit
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  • 3
    Digitale Medien
    Digitale Medien
    Cloning, sequence and expression of the gene coding for rhamnogalacturonase ofAspergillus aculeatus; a novel pectinolytic enzyme (1995)
    Springer
    Applied microbiology and biotechnology 43 (1995), S. 861-870 
    Details
    ISSN: 1432-0614
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie , Werkstoffwissenschaften, Fertigungsverfahren, Fertigung
    Notizen: Abstract Rhamnogalacturonase was purified from culture filtrate ofAspergillus aculeatus after growth in medium with sugar-beet pulp as carbon source. Purified protein was used to raise antibodies in mice and with the antiserum obtained a gene coding for rhamnogalacturonase (rhgA) was isolated from a λ cDNA expression library. The clonedrhgA gene has an open-reading frame of 1320 base pairs encoding a protein of 440 amino acids with a predicted molecular mass of 45 962 Da. The protein contains a potential signal peptidase cleavage site behind Gly-18 and three potential sites forN-glycosylation. Limited homology withA. niger polygalacturonase amino acid sequences is found. A genomic clone ofrhgA was isolated from a recombinant phage λ genomic library. Comparison of the genomic and cDNA sequences revealed that the coding region of the gene is interrupted by three introns. Furthermore, amino acid sequences of four different peptides, derived from purifiedA. aculeatus rhamnogalacturonase, were also found in the deduced amino acid sequence ofrhgA.A. aculeatus strains overexpressing rhamnogalacturonase were obtained by cotransformation using either theA. niger pyrA gene or theA. aculeatus pyr A gene as selection marker. For expression of rhamnogalacturonase inA. awamori theA. awamori pyrA gene was used as selection marker. Degradation patterns of modified hairy regions, determined by HPLC, show the recombinant rhamnogalacturonase to be active, and the enzyme was found to have a positive effect in the apple hot-mash liquefaction process.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF02431920
    Bibliothek Standort Signatur Band/Heft/Jahr Verfügbarkeit
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    Artikel (Nationallizenzen)
    Volltext
  • 4
    Digitale Medien
    Digitale Medien
    A novel strategy for the isolation of defined pyrG mutants and the development of a site-specific integration system for Aspergillus awamori (1995)
    Springer
    Current genetics 27 (1995), S. 536-540 
    Details
    ISSN: 1432-0983
    Schlagwort(e): Recombinant DNA ; Filamentous fungi ; 5-fluoro-orotic acid ; Homologous transformation system
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract A homologous gene transfer system for Aspergillus awamori for site-specific integration is described, based on two components. First, a defined A. awamori pyrG mutant strain constructed by a selection strategy for gene-replacement in fungi. Second, a vector with a homologous pyrG selection marker containing a defined mutation at a site different from that of the mutations in the pyrG gene of the defined mutant strain. Defined mutation in the A. awamori pyrG gene, isolated from a genomic library by heterologous hybridisation with the A. niger pyrG gene as a probe, were introduced by specifically altering sequences at restriction sites in the coding region of the gene. After transformation of the A. awamori wild-type strain with vectors containing these mutated pyrG genes, and selection for 5-fluoro-orotic acid resistance (5-FOAR), on the average 60% of the 5-FOAR colonies originated from replacement of the wild-type pyrG gene by the mutated pyrG allele. After transformation of a mutant strain, carrying a mutation near the 5′ end of the pyrG gene with vectors containing a mutation near the 3′ end of the pyrG gene, 35% of the resulting transformants contained one copy of the vector at the pyrG locus.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF00314444
    Bibliothek Standort Signatur Band/Heft/Jahr Verfügbarkeit
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    Artikel (Nationallizenzen)
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