The development of functionalized nanomaterial biosynthesis processes is important to expand many cutting-edge nanomaterial application areas. However, unclear synthesis mechanisms and low synthesis efficiency under various chemical stresses have limited the use of these biomaterials. Deinococcus radiodurans is an extreme bacterium well known for its exceptional resistance to radiation oxidants and electrophilic agents. This extremophile, which possesses a spontaneous self-assembled surface-layer (S-layer), has been an optimal model organism to study microbial nanomaterial biotemplates and biosynthesis under various stresses. This review summarizes the S-layers from D. radiodurans as an excellent biotemplate for various pre-synthesized nanomaterials and multiple applications, and highlights recent progresses about the biosynthesis of functionalized gold nanoparticles (AuNPs), silver nanoparticles (AgNPs), as well as gold and silver bimetallic nanoparticles using D. radiodurans. Their formation mechanisms, properties, and applications are discussed and summarized to provide significant insights into the design or modification of functionalized nanomaterials via natural materials. Grand challenges and future directions to realize the multifunctional applications of these nanomaterials are highlighted for a better understanding of their biosynthesis mechanisms and functionalized modifications. © 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.