De novo design of small molecule–regulated protein oligomers

Small molecule–regulated protein oligomerization provides a powerful mechanism for manipulating biological processes by controlling protein proximity with high temporal precision. However, such systems only rarely exist in nature and remain a substantial challenge for de novo design. In this work, we describe a computational method for designing protein homooligomers whose assembly is regulated by small-molecule ligands with matching symmetry. We designed protein homotrimers regulated by the Food and Drug Administration (FDA)–approved drug amantadine and further designed amantadine-responsive heterodimers and heterotrimers. Biophysical characterization confirmed their amantadine-dependent assembly, and their crystal structures closely matched the design models. We demonstrated their broad applicability in controlling protein localization, membraneless condensate formation, and gene expression. Our approach opens new avenues for designing small molecule–responsive proteins and expands the chemogenetic toolkit for manipulating complex biological processes.