Heterologous Expression of Plant Cell Wall Degrading Enzymes for Effective Production of Cellulosic Biofuels

Table 2

Factors to consider in plant-based expression of plant cell wall degrading enzymes.

Category	Exemplary factors associated with a common strategy
Expression type	Selection between stable transformation in a transgenic plant and transient expression in a wildtype plant
Host plant	Composition of biomass; Ease of DNA cloning; Productivity of a plant crop; Selection between monocot and dicot plants
Gene	Development of fusion protein; Function and properties of enzyme for example, Thermal stability; Subcellular protein localization using transit peptide; Synthetic gene design for example, Codon/UTR optimization; Use of plant cell wall modifying enzyme
Expression vector	Development of viral expression system for transient gene expression; Easy selection markers for stably transformed plants; Optimal promoter; RBS optimization for example, Kozak's context sequence for monocot; Selection between constitutive and inducible systems
Functional analysis	Change of enzyme stability with time; Cleavage of holoenzyme for example, Truncated enzyme; Comparison of plant produced enzyme with E. coli produced enzyme; Effect of plant crude extract on enzyme stability; Expression of multiple enzymes; Microscopic analysis of protein localization; Physical properties for example, Activity optima, thermal stability; Posttranslational modification for example, Glycosylation; Purification process
Phenotype analysis of transgenic plant	Compositional and structural change in transgenic plant; Deleterious phenotype change for example, Fertile, pale leaves; Location of protein accumulation for example, High level accumulation in leaf or stem; New beneficial traits for example, Resistance to pathogens
Biomass hydrolysis	Application of enzyme cocktails; Composition and property of cellulosic biomass; Induction of in-planta hydrolysis; Optimal combination of enzymes; Resistance of enzymes to pretreatment process