Proteomic Profiling of the Secretome of Trichoderma reesei

• Published in: Methods in molecular biology (Clifton, N.J.) Vol. 2234; p. 237
• Main Authors: Ngan, So Fong Cam, Sze, Siu Kwan
• Format: Journal Article
• Language: English
• Published: United States 2021
• Subjects: Cells, Cultured; Chromatography, Liquid; Data Analysis; Fungal Proteins - metabolism; Hypocreales - metabolism; Peptides - metabolism; Proteomics - methods; Staining and Labeling; Tandem Mass Spectrometry; LC-MS/MS; Secretome; Lignocellulose; Cellulolytic enzymes; Trichoderma reesei; Proteomics
• ISSN: 1940-6029
• DOI: 10.1007/978-1-0716-1048-0_17

Trichoderma reesei (T. reesei) is the workhorse for the production of industrial cellulolytic enzyme cocktails for cellulose hydrolysis. However, the current industrial process using enzyme cocktails is not efficient enough for the cost-effective generation of cellulosic sugar. Here, we describe a protocol for the application of a state-of-the-art LC-MS/MS-based proteomics method for studying the T. reesei secretome. A protein-free minimal chemically defined cell culture medium must be used for a successful secretome analysis. A lignocellulose substrate can be added to this minimal medium to stimulate the fungal secretion of enzymes specific to that substrate. The secretory proteins in the conditioned medium can be purified for quantitative proteomics profiling. T. reesei secretes several hundred enzymes including cellulases, hemicellulases, pectinases, proteases, oxidoreductases, and many putative proteins when it is stimulated with lignocellulose. By combining an understanding of the basic biomass hydrolytic mechanisms with the discovery of novel enzymes, more effective enzyme cocktails can be designed for a sustainable biochemical-based biorefinery.