The conversion of lignocellulosic biomass into biofuels or biochemicals typically involves a pretreatment process followed by the enzyme-catalyzed hydrolysis of cellulose and hemicellulose components to fermentable sugars. Many factors can contribute to the recalcitrance of biomass, e.g., the lignin content and structure, crystallinity of cellulose, degree of fiber polymerization, and hemicellulose content, among others. However, nonproductive binding between cellulase and lignin is the factor with the greatest impact on enzymatic hydrolysis. To reduce the nonproductive adsorption of enzymes on lignin and improve the efficiency of enzymatic hydrolysis, this review comprehensively summarized the progress that has been made in understanding the interactions between lignin and enzymes. Firstly, the effects of pretreatment techniques on lignin content and enzymatic hydrolysis were reviewed. The effects of lignin content and functional groups on enzymatic hydrolysis were then summarized. Methods for the preparation and characterization of lignin films were assessed. Finally, the methods applied to characterize the interactions between lignin and cellulase were reviewed, and methods for decreasing the nonproductive binding of enzymes to lignin were discussed. This review provides an overview of the current understanding of how lignin hinders the enzymatic hydrolysis of lignocellulosic biomass, and provides a theoretical basis for the development of more economical and effective methods and additives to reduce the interaction of lignin and enzymes to improve the efficiency of enzymatic hydrolysis.