Currently, gasoline and other nonrenewable fossil fuels encompass most of the fuel industry. However, these fuels are causing irreversible environmental damage, and the biofuel industry rose as an alternative. Biofuels utilize biomass as their energy source and are a greener and more sustainable solution. Yet, the chemical reactions previously conducted to produce biofuels have been inefficient and costly due to the enzyme’s instability and lack of reusability. Enzymes are natural biological molecules that can catalyze chemical reactions, and enzyme immobilization is a technique that fixes an enzyme to a support to improve efficiency. Enzyme immobilization comes in the form of many unique approaches, such as encapsulation, entrapment, covalent bonding, adsorption, and cross-linking. Each approach provides distinct advantages and disadvantages and has applications in the biofuel industry. Notwithstanding the different approaches, various supports for enzyme immobilization exist and include organic supports, inorganic supports, natural polymers, synthetic polymers, and nanomaterials. This chapter emphasizes nanomaterials, as nanomaterials are supports with pronounced potential. These immobilization processes and nanomaterials as supports culminate in the production of biofuels like bioethanol, biogas, biokerosene, biohydrogen, biodiesel, and biobutanol. The reactions producing biofuels are then quantitatively compared using kinetic and thermodynamic parameters. In closing, this chapter aims to spread awareness of the benefits of using enzyme immobilization to increase biofuel production, thus creating a sustainable solution for the future.