SAMHD1 is a GTP-activated nonspecific dNTP triphosphohydrolase that depletes dNTP pools in resting CD4+ T cells and macrophages and effectively restricts infection by HIV-1. We have designed a nonsubstrate dUTP analogue with a methylene bridge connecting the a phosphate and 5' carbon that potently inhibits SAMHD1. Although pppCH(2)dU shows apparent competitive inhibition, it acts by a surprising allosteric mechanism that destabilizes active enzyme tetramer.