title: The effects of bioprocess parameters on extracellular proteases in a recombinant Aspergillus niger B1-D.
abstract: Although host proteases are often considered to have a negative impact upon heterologous protein production by filamentous fungi, relatively little is known about the pattern of their appearance in recombinant fungal bioprocesses. In the present study, we investigated extracellular proteases from a filamentous fungus, Aspergillus niger B1-D, genetically modified to secrete hen egg white lysozyme (HEWL). Our findings indicate that extracellular protease activity is only detected after the carbon source is completely utilised in batch cultures. The proteases are predominantly acid proteases and have optimal temperature for activity at around 45 degrees C. Their activity could be partially inhibited by protease inhibitors, indicating the existence of at least four kinds of proteases in these culture fluids, aspartic-, serine-, cysteine-, and metallo-proteases. Oxygen enrichment does not have any noticeable effects on extracellular protease activity except that the onset of protease activity appears earlier in oxygen enrichment runs. Oxygen enrichment stimulates HEWL production substantially, and we propose that it is related to fungal morphology. Thermal stress imposed by raising process temperature (from 25 to 30 and 35 degrees C) in early exponential phase, led to appearance of protease activity in the medium following the heat shock. Continued cultivation at high temperatures significantly reduced HEWL production, which was associated with increased activity of the extracellular proteases in these cultures.
authors: Strathclyde Fermentation Centre, Strathclyde Institute of Pharmacy and Biomedical Sciences, University of Strathclyde, 204 George Street, Glasgow, G1 1XW, UK.
authors: Qiang
authors: Q
authors: Li
authors: Linda M
authors: LM
authors: Harvey
authors: Brian
authors: B
authors: McNeil
mesh_terms: D016282:Aspartic Acid Endopeptidases / Q000378:metabolism; D001234:Aspergillus niger / Q000201:enzymology* / Q000235:genetics; D001426:Bacterial Proteins / Q000378:metabolism*; D002244:Carbon / Q000378:metabolism; D003546:Cysteine Endopeptidases / Q000378:metabolism; D045726:Metalloproteases / Q000378:metabolism; D009113:Muramidase / Q000096:biosynthesis / Q000235:genetics; D010100:Oxygen / Q000378:metabolism; D010447:Peptide Hydrolases / Q000378:metabolism*; D011480:Protease Inhibitors / Q000494:pharmacology; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D012697:Serine Endopeptidases / Q000378:metabolism; D013696:Temperature
chemical_list: D001426:Bacterial Proteins; D011480:Protease Inhibitors; D011994:Recombinant Proteins; D002244:Carbon; C421864:hen egg lysozyme; D009113:Muramidase; D045726:Metalloproteases; D010447:Peptide Hydrolases; D012697:Serine Endopeptidases; D003546:Cysteine Endopeptidases; D016282:Aspartic Acid Endopeptidases; D010100:Oxygen