Molecular and Genetic Strategies for Enhanced Production of Heterologous Lignocellulosic Enzymes

Abstract

A desire to move away from fossil fuels toward more sustainable, renewable, or carbon-neutral sources has motivated the search for alternative feedstocks. Lignocellulosic biomass is an attractive option due to its abundance, low cost, and high sugar content. In nature, the breakdown of lignocellulosic biomass is a sequential process involving the collective contribution of numerous microorganisms and enzymes. In biorefineries, however, the high cost and limited availability of the enzyme cocktails required for efficient biomass deconstruction are significant barriers to the adoption of lignocellulosic biomass as a feedstock to produce biofuels and other value-added products. Filamentous fungi have an astounding capacity to secrete digestive enzymes into their extracellular environment through the presence of efficient systems for the transcription, translation, folding, posttranslational modification, and secretion of polypeptides. These features have led filamentous fungi to be exploited for the manufacturing of homologous and heterologous proteins for the chemical, pharmaceutical, and biofuel industries. Total homologous (native) enzyme expression can reach about 10–100 g/L, but heterologous enzyme production is hampered by much lower concentration. Efficient heterologous protein production often requires multilevel optimization strategies for both the protein and the expression host as factors limiting production are often protein-specific and bottlenecks can exist at any stage from transcription and translation to folding and secretion. Here, we review the molecular and genetic strategies used for the particularly challenging case of heterologous enzyme production in filamentous fungi, with an emphasis on the enzymes involved in the conversion of lignocellulosic biomass. The development of efficient expression systems and hosts to identify, characterize, and enhance the expression levels, yields, and activities of heterologous lignocellulosic enzymes is critical to the successful adoption of lignocellulosic biomass into the new bio-economy.