Enzyme stability and activity are pivotal factors for their implementation in different industrial applications. Enzyme supramolecular engineering relies on the fabrication of a tailor-made enzyme nano-environment to ensure enzyme stability without impairing activity. Cyclodextrins (CDs), cyclic oligomers of glucose, act as protein chaperones and stabilize, upon interaction with hydrophobic amino acid residues exposed at the protein surface, its three-dimensional structure. When used to build an organosilica layer shielding an enzyme, they enhance the protective effect of this layer. In the present study, we systematically assessed the protective effects of three organosilane derivatives based on ɑ-, β- and γ-CDs. A model lipase enzyme was immobilized at the surface of silica nanoparticles and shielded in an organosilica layer containing these organosilanes. Besides layer thickness optimization, the effect of different stressors (i. e., temperature, SDS, urea) was tested. Our results showed that organosilica layers produced with CDs improve enzyme thermal stability. They also support enzyme refolding after denaturation under chaotic conditions. Additionally, we demonstrated that the protective effect of the smallest CD derivative tested, namely ɑ-CD, surpassed the other macrocycles studied for conferring the immobilized enzyme with higher resistance to stress conditions. This protection strategy was also applied to a thermostable β-galactosidase enzyme.