A defined level of protein disulfide isomerase expression is required for optimal secretion of thaumatin by Aspegillus awamori.
Thaumatin, a 22-kDa protein containing eight disulfide bonds, is secreted by the filamentous fungus Aspergillus awamori at levels which are dependent upon the extent of overexpression of protein disulfide isomerase (PDIA). Additional copies of the PDIA-encoding gene pdiA were introduced into a strain of A. awamori that expresses a cassette encoding thaumatin. Transformants with different levels of pdiA mRNA and measured PDIA levels were chosen for examination of the impact that PDIA levels had on thaumatin secretion. The secretion of two native proteins, alpha-amylase and acid phosphatase, was also examined in relation to varying levels of PDIA. Over a range of PDIA levels of 1-8, relative to the native level in strains with just one copy of the pdiA gene, the fraction of alpha-amylase and acid phosphatase in the total secreted protein was unaffected. In contrast, a peak level of thaumatin, about 5-fold higher than in the strain with one copy of pdiA, was found in strains with a relative PDIA level of between two and four. Improved thaumatin production was confirmed in 5-1 fermenters using a strain of A. awamori with six pdiA gene copies, containing 3.2-fold higher levels of PDIA than wild-type strains.
Moralejo
F J
FJ
Institute of Biotechnology INBIOTEC, Science Park of León, Spain.
Watson
A J
AJ
Jeenes
D J
DJ
Archer
D B
DB
Martín
J F
JF
D000135:Acid Phosphatase / Q000378:metabolism; D001230:Aspergillus / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D005285:Fermentation; D005656:Fungal Proteins / Q000378:metabolism; D005784:Gene Amplification; D015971:Gene Expression Regulation, Enzymologic; D015966:Gene Expression Regulation, Fungal; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism*; D019704:Protein Disulfide-Isomerases / Q000235:genetics* / Q000378:metabolism; D013549:Sweetening Agents; D014170:Transformation, Genetic; D000516:alpha-Amylases / Q000378:metabolism
D005656:Fungal Proteins; D010940:Plant Proteins; D013549:Sweetening Agents; C003427:thaumatin protein, plant; D000135:Acid Phosphatase; D000516:alpha-Amylases; D019704:Protein Disulfide-Isomerases