@article{1ed3186e1eab4d77ad2df081c5f434f9,
title = "Tsg101 chaperone function revealed by HIV-1 assembly inhibitors",
abstract = "HIV-1 replication requires Tsg101, a component of cellular endosomal sorting complex required for transport (ESCRT) machinery. Tsg101 possesses an ubiquitin (Ub) E2 variant (UEV) domain with a pocket that can bind PT/SAP motifs and another pocket that can bind Ub. The PTAP motif in the viral structural precursor polyprotein, Gag, allows the recruitment of Tsg101 and other ESCRTs to virus assembly sites where they mediate budding. It is not known how or even whether the UEV Ub binding function contributes to virus production. Here, we report that disruption of UEV Ub binding by commonly used drugs arrests assembly at an early step distinct from the late stage involving PTAP binding disruption. NMR reveals that the drugs form a covalent adduct near the Ub-binding pocket leading to the disruption of Ub, but not PTAP binding. We conclude that the Ub-binding pocket has a chaperone function involved in bud initiation.",
author = "Madeleine Strickland and Ehrlich, {Lorna S.} and Susan Watanabe and Mahfuz Khan and Strub, {Marie Paule} and Luan, {Chi Hao} and Powell, {Michael D.} and Jonathan Leis and Nico Tjandra and Carter, {Carol A.}",
note = "Funding Information: We thank Dr. Fadila Bouamr for providing the construct encoding the HIV-1 Gag-Herpes Simplex virus deubiquitinating enzyme chimeric protein; Sara F. Dunne of the Northwestern U High Throughput Screening (HTS) Facility for assistance with high-throughput screening; Susan van Horn (Stony Brook University Central Microscopy Core Facility) for electron microscopy services. We also thank Dr. Duck-Yeon Lee of the Biochemistry Core Facility and Yi He of the Laboratory of Structural Biophysics at the National Heart, Lung and Blood Institute (NHLBI) of the National Institutes of Health (NIH) for expertise and advice regarding LC−MS and for the growth of E. coli for production of isotopically-labeled Tsg101, respectively. We also acknowledge Dr. Guil-lermo A. Bermejo, Dr. Yong-Sok Lee, and Dr. Charles D. Schwieters at the Center for Information Technology at the NIH for help with preparing structure files for N16 and for help with structure calculations using Xplor-NIH. These investigations were supported by funds from the NIH National Institute of General Medical Sciences (NIGMS) R01 GM111028 and from the NHLBI (U01HL127522) and the New York State Department of Economic Development (C140151) and by the Center for Biotechnology, Stony Brook University to C.A.C.; NHLBI (U01HL127522) and NHLBI Intramural Research Programs to N.T.; Northwestern Memorial Hospital Dixon Innovation Grant and Chicago Biomedical Consortium/HTS Supplemental Grant to J.L. and C-H.L.; Lurie Cancer Center grant #P30CA060553 to C-H.L.; RCMI grant G12MD007602 and R21 NS105577 to M.D.P. The work is related to pending patent applications which name some of the authors as inventors. Publisher Copyright: {\textcopyright} 2017 The Author(s).",
year = "2017",
month = dec,
day = "1",
doi = "10.1038/s41467-017-01426-2",
language = "English (US)",
volume = "8",
journal = "Nature Communications",
issn = "2041-1723",
publisher = "Nature Publishing Group",
number = "1",
}
