TY - JOUR T1 - Strategies to target SARS-CoV-2 entry and infection using dual mechanisms of inhibition by acidification inhibitors. JF - PLoS Pathog Y1 - 2021 A1 - Prabhakara, Chaitra A1 - Godbole, Rashmi A1 - Sil, Parijat A1 - Jahnavi, Sowmya A1 - Gulzar, Shah-E-Jahan A1 - van Zanten, Thomas S A1 - Sheth, Dhruv A1 - Subhash, Neeraja A1 - Chandra, Anchal A1 - Shivaraj, Akshatha A1 - Panikulam, Patricia A1 - U, Ibrahim A1 - Nuthakki, Vijay Kumar A1 - Puthiyapurayil, Theja Parassini A1 - Ahmed, Riyaz A1 - Najar, Ashaq Hussain A1 - Lingamallu, Sai Manoz A1 - Das, Snigdhadev A1 - Mahajan, Bhagyashri A1 - Vemula, Praveen A1 - Bharate, Sandip B A1 - Singh, Parvinder Pal A1 - Vishwakarma, Ram A1 - Guha, Arjun A1 - Sundaramurthy, Varadharajan A1 - Mayor, Satyajit KW - Ammonium Chloride KW - Angiotensin-Converting Enzyme 2 KW - Animals KW - Antiviral Agents KW - Cell Line KW - Chlorocebus aethiops KW - Chloroquine KW - Clathrin KW - COVID-19 KW - Drug Synergism KW - Endocytosis KW - Endosomes KW - Humans KW - Hydrogen-Ion Concentration KW - Hydroxychloroquine KW - Macrolides KW - Niclosamide KW - Protein Binding KW - Protein Domains KW - SARS-CoV-2 KW - Spike Glycoprotein, Coronavirus KW - Vero Cells KW - Virus Internalization AB -

Many viruses utilize the host endo-lysosomal network for infection. Tracing the endocytic itinerary of SARS-CoV-2 can provide insights into viral trafficking and aid in designing new therapeutic strategies. Here, we demonstrate that the receptor binding domain (RBD) of SARS-CoV-2 spike protein is internalized via the pH-dependent CLIC/GEEC (CG) endocytic pathway in human gastric-adenocarcinoma (AGS) cells expressing undetectable levels of ACE2. Ectopic expression of ACE2 (AGS-ACE2) results in RBD traffic via both CG and clathrin-mediated endocytosis. Endosomal acidification inhibitors like BafilomycinA1 and NH4Cl, which inhibit the CG pathway, reduce the uptake of RBD and impede Spike-pseudoviral infection in both AGS and AGS-ACE2 cells. The inhibition by BafilomycinA1 was found to be distinct from Chloroquine which neither affects RBD uptake nor alters endosomal pH, yet attenuates Spike-pseudovirus entry. By screening a subset of FDA-approved inhibitors for functionality similar to BafilomycinA1, we identified Niclosamide as a SARS-CoV-2 entry inhibitor. Further validation using a clinical isolate of SARS-CoV-2 in AGS-ACE2 and Vero cells confirmed its antiviral effect. We propose that Niclosamide, and other drugs which neutralize endosomal pH as well as inhibit the endocytic uptake, could provide broader applicability in subverting infection of viruses entering host cells via a pH-dependent endocytic pathway.

VL - 17 IS - 7 ER - TY - JOUR T1 - Metal substitutions incarbonic anhydrase: a halide ion probe study. JF - Biochem Biophys Res Commun Y1 - 1975 A1 - Smith, R J A1 - Bryant, R G KW - Animals KW - Binding Sites KW - Cadmium KW - Carbonic Anhydrases KW - Cattle KW - Humans KW - Hydrogen-Ion Concentration KW - Magnetic Resonance Spectroscopy KW - Mercury KW - Protein Binding KW - Protein Conformation KW - Zinc VL - 66 IS - 4 ER -