Cloning and characterization of the abfB gene coding for the major α-l-arabinofuranosidase (ABF B) of Aspergillus niger

Michel J.A. Flipphi, Margreet van Heuvel, Peter van der Veen, Jaap Visser, Leo H. de Graaff

Research output: Contribution to journalArticle

58 Citations (Scopus)

Abstract

Based on amino-acid sequence data from Aspergillus niger α-l-arabinofuranosidase B (ABF B), and cyanogen bromide fragments derived thereof, deoxyoligonucleotide mixtures were designed to the employed as primers in a polymerase chain reaction (PCR) on A. niger genomic DNA. This resulted in amplification of three related PCR products. The abfB gene encoding ABF B was isolated from a genomic library using such an amplification product as a probe. A 5.1-kb BamHI fragment was subcloned to result in plasmid pIM991. Upon introduction by co-transformation into both A. niger and A. nidulans uridine auxotrophic strains, pIM991 was shown to contain the functional gene since prototrophic transformants overproduced ABF B upon growth on the inducing carbon source sugar beet pulp. A plate assay was developed enabling quick selection of ABF B-overproducing transformants. The sequence of a 4122-bp long BamHI/SstI fragment was determined. The abfB gene does not contain introns and codes for a protein of 499 amino acids. The mature ABF B, 481 amino acids in length, has a deduced molecular weight of 50.7 kDa. A. niger abfB is the first eukaryotic gene encoding an ABF to be characterized.

Original languageEnglish
Pages (from-to)525-532
Number of pages8
JournalCurrent Genetics
Volume24
Issue number6
DOIs
Publication statusPublished - Dec 1 1993

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Keywords

  • Aspergillus nidulans
  • Aspergillus niger
  • DNA sequence
  • Fungal transformation
  • Gene structure
  • Heterologous expression
  • Overexpression
  • α-l-Arabinofuranosidase

ASJC Scopus subject areas

  • Genetics

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