Diverse virus families have evolved to exploit the acidification of endosomal compartments to gain entry into cells. We describe a supramolecular approach for selectively targeting and inhibiting viral infections through this central biochemical pathway. Using adenovirus as a model non-enveloped virus, we have determined that an eight-residue cyclic D,L-alpha-peptide, selected from a directed combinatorial library, can specifically prevent the development of low pH in endocytic vesicles, arrest the escape of virions from the endosome, and abrogate adenovirus infection without an apparent adverse effect on cell viability. The likely generality of this approach against other pH-dependent viral infections is supported by the inhibition of type-A influenza virus escape from endosomes in the presence of the same peptide. Our studies suggest that self-assembling cyclic D,L-alpha-peptides hold considerable potential as a new rational supramolecular approach toward the design and discovery of broad-spectrum antiviral agents.