The ubiquitin E2 variant domain of TSG101 (TSG101-UEV) plays a pivotal role in protein sorting and virus budding by recognizing PTAP motifs within ubiquitinated proteins. Disruption of TSG101-UEV/PTAP...

The ubiquitin E2 variant domain of TSG101 (TSG101-UEV) plays a pivotal role in protein sorting and v

Plasmids expressing the fluorescently tagged proteins ALIX, TSG101, CHMP2A, VPS4A, and VPS4B were described previously39. Plasmids expressing the doxycycline inducible E228Q-VPS4A-HA, mCherry-Tomm20-N10, and pCHAC-mt-mKeima were obtained from Addgene (#200087, #55146, #72342). pLKO.1-puromycin plasmids expressing shRNA targeting to TSG101, CHMP3, VPS4B,...

The ubiquitin E2 variant domain of TSG101 (TSG101-UEV) plays a pivotal role in protein sorting and v

Characterization of a family of covalent prazole derivative adducts with the Tsg101 UEV domain finds roles for bulk and flexibility in antiviral efficacy and cytotoxicity, while opening avenues for further development of prazole-based antiviral therapeutics that target Tsg101. Abstract Here, an in vitro characterization of a family of prazole derivatives that covalently bind to the C73 site on Tsg101 and assay their ability to inhibit viral particle production is presented. Structurally, increased steric bulk on the 4-pyridyl of the prazole expands the prazole site on the UEV domain toward the β-hairpin in the Ub-binding site and is coupled to increased inhibition of virus-like particle production in HIV-1. Increased bulk also increased toxicity, which is alleviated by increasing flexibility. Further, the formation of a novel secondary Tsg101 adduct for several of the tested compounds and the commercial drug lansoprazole. The secondary adduct involved the loss of the 4-pyridyl substituent to form an irreversible species, with implications for increasing the half-life of the active species or its specificity toward Tsg101 UEV. It is also determined that sulfide derivatives display effective viral inhibition, presumably through cellular sulfoxidation, allowing for delayed conversion within the cellular environment, and identify SARS-COV-2 as a target of prazole inhibition. These results open multiple avenues for the design of prazole derivatives for antiviral applications.