Time course analysis of the extracellular proteome of Aspergillus nidulans growing on sorghum stover

Abstract

Background: Fungi are important players in the turnover of plant biomass because they produce a broad range of degradative enzymes. Aspergillus nidulans, a well-studied saprophyte and close homologue to industrially important species such as A. niger and A. oryzae, was selected for this study.

Results: A. nidulans was grown on sorghum stover under solid-state culture conditions for 1, 2, 3, 5, 7 and 14 days. Based on analysis of chitin content, A. nidulans grew to be 4-5% of the total biomass in the culture after 2 days and then maintained a steady state of 4% of the total biomass for the next 12 days. A hyphal mat developed on the surface of the sorghum by day one and as seen by scanning electron microscopy the hyphae enmeshed the sorghum particles by day 5. After 14 days hyphae had penetrated the entire sorghum slurry. Analysis (1-D PAGE LC-MS/MS) of the secretome of A. nidulans, and analysis of the breakdown products from the sorghum stover showed a wide range of enzymes secreted. A total of 294 extracellular proteins were identified with hemicellulases, cellulases, polygalacturonases, chitinases, esterases and lipases predominating the secretome. Time course analysis revealed a total of 196, 166, 172 and 182 proteins on day 1, 3, 7 and 14 respectively. The fungus used 20% of the xylan and cellulose by day 7 and 30% by day 14. Cellobiose dehydrogenase, feruloyl esterases, and CAZy family 61 endoglucanases, all of which are thought to reduce the recalcitrance of biomass to hydrolysis, were found in high abundance.

Conclusions: Our results show that A. nidulans secretes a wide array of enzymes to degrade the major polysaccharides and lipids (but probably not lignin) by 1 day of growth on sorghum. The data suggests simultaneous breakdown of hemicellulose, cellulose and pectin. Despite secretion of most of the enzymes on day 1, changes in the relative abundances of enzymes over the time course indicates that the set of enzymes secreted is tailored to the specific substrates available. Our findings reveal that A. nidulans is capable of degrading the major polysaccharides in sorghum without any chemical pre-treatment.