Metabolically independent and accurately adjustable Aspergillus sp. expression system

Area of Science:

• Biotechnology
• Molecular Biology
• Mycology

Background:

• Filamentous fungi, like Aspergillus species, are widely used for producing extracellular proteins in food and pharmaceutical industries.
• Current expression systems in Aspergillus rely on constitutive or inducible promoters linked to metabolic pathways.
• Existing systems face limitations in control and media requirements.

Purpose of the Study:

• To develop and characterize a novel, tightly regulated gene expression system for Aspergillus nidulans and Aspergillus niger.
• To utilize the human estrogen receptor transcriptional activation by estrogenic substances for inducible gene expression.
• To establish a system independent of metabolic signals, enabling use with low-cost media.

Main Methods:

• A novel expression system was designed for Aspergillus species, employing the human estrogen receptor for transcriptional activation.
• The system integrates positive and negative regulatory elements within the promoter to control reporter gene expression.
• Induction was achieved using estrogenic substances, with dose-response and expression levels monitored.

Main Results:

• The novel system demonstrated tight 'off' status with rapid induction (within minutes) and high expression levels (up to 15% of total cell protein after 8 hours).
• Expression showed a linear dose-response to the inducer, functioning independently of metabolic signals and allowing use of complex media.
• Aspergillus species exhibited high sensitivity to estrogenic substances, with low-cost inducers effective at picomolar concentrations.

Conclusions:

• A novel, estrogen-inducible gene expression system for Aspergillus species has been successfully developed.
• This system offers precise control, high protein yield, and flexibility in media composition.
• The high sensitivity of Aspergillus to estrogens suggests potential applications in detecting environmental xenoestrogens.