Isolation and characterization of mutants of Aspergillus niger deficient in extracellular proteases

Mol Gen Genet. 1992 Aug;234(2):332-6. doi: 10.1007/BF00283855.

Abstract

In the present study, the extracellular protease activity in a strain of the filamentous fungus Aspergillus niger was investigated and mutant strains deficient in the production of extracellular proteases were isolated. The major protease, which is responsible for 80-85% of the total activity, is aspergillopepsin A, a protein of ca. 43 kDa, the activity of which is inhibited by pepstatin. In addition, a second protease, aspergillopepsin B, is produced, which is much less sensitive to inhibition by pepstatin. Several protease-deficient mutants were obtained by in vivo UV mutagenesis. In addition, a mutant lacking aspergillopepsin A was constructed by an in vitro gene replacement strategy. In this mutant, AB1.1, the entire coding region of the gene for aspergillopepsin A (pepA) is deleted. In three UV-induced mutants, aspergillopepsin A is also missing. One of these mutants, AB1.18, is mutated in the pepA gene, which is located on chromosome I. One of the other mutants, AB1.13, which has only 1-2% of the extracellular protease activity in the parent strain, is deficient in both aspergillopepsin A and aspergillopepsin B. The mutation involved, prt-13, has been localized to chromosome VI, and is probably a mutation in a regulatory gene. Another mutation involved in loss of protease function, prt-39, is located on chromosome VIII. Degradation of various heterologous proteins in culture media of the mutants is reduced but, even in strain AB1.13, not completely abolished.

MeSH terms

  • Aspartic Acid Endopeptidases / genetics*
  • Aspartic Acid Endopeptidases / metabolism
  • Aspergillus niger / enzymology*
  • Aspergillus niger / genetics
  • Aspergillus niger / isolation & purification
  • Blotting, Western
  • Fungal Proteins / genetics*
  • Fungal Proteins / metabolism
  • Mutagenesis / genetics

Substances

  • Fungal Proteins
  • Aspartic Acid Endopeptidases
  • aspergillopepsin I