Ribosome-inactivating proteins (RIPs) are N-glycosidases. They depurinate A-4324 in rat 28S ribosomal RNA in the conserved α-sarcin/ricin loop (α-SRL) and cease protein synthesis. Our group has shown that the internal peptide of the maize RIP precursor reduced the anti-HIV activity of the protein in infected macaque peripheral blood mononuclear cells (PBMC) and SHIV 89.6-infected Chinese rhesus macaque. We made use of the switch-on mechanism of maize RIP to incorporate HIV-1 protease recognition sequences to its internal inactivation region. Upon activation of this engineered maize RIP by HIV-1 protease in HIV-infected cells, the N-glycosidase activity and inhibitory effect on p24 antigen production in vitro and in infected human T cells were enhanced. This switch-on mechanism can also be applied to ricin A chain (RTA). RTA variants with HIV-1 protease recognition sequence at the C-terminus can be cleaved both in vitro and in HIV-infected cells. Furthermore, its antiviral effect was enhanced and the cytotoxicity towards uninfected cells was reduced. Our study provides a platform technology in creating protein toxin derivatives with increased pathogen-specific cytotoxicity.