Display omitted •Interactome represents a whole set of interactions during lignocellulose breakdown.•Fungus-substrate interactome drives enzyme expression.•Enzyme-enzyme interactome involves synergism and LMW molecules exchanging.•Interactome can be driven to enhance biomass valorization.•Interactome can unlock the full potential of lignocellulosic components. Biomass represents an abundant and inexpensive source of sugars and aromatic compounds that can be used as raw materials for conversion into value-added bioproducts. Filamentous fungi are sources of plant cell wall degrading enzymes in nature. Understanding the interactions between enzymes is crucial for optimizing biomass degradation processes. Herein, the concept of the interactome is presented as a holistic approach that depicts the interactions among enzymes, substrates, metabolites, and inhibitors. The interactome encompasses several stages of biomass degradation, starting with the sensing of the substrate and the subsequent synthesis of hydrolytic and oxidative enzymes (fungus-substrate interaction). Enzyme-enzyme interactions are exemplified in the complex processes of lignocellulosic biomass degradation. The enzyme-substrate-metabolite-inhibitor interaction also provides a better understanding of biomass conversion, allowing bioproduct production from recalcitrant agro-industrial residues, thus bringing greater value to residual biomass. Finally, technological applications are presented for optimizing the interactome at various levels.