Compounds derived from Humulus lupulus inhibit SARS-CoV-2 papain-like protease and virus replication.

BACKGROUND: Selected natural compounds exhibit very good antiviral properties. Especially, the medicinal plant Humulus lupulus (hop) contains several secondary plant metabolites some of which have previously shown antiviral activities. Among them, the prenylated chalcone xanthohumol (XN) demonstrated to be a potent inhibitor of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) main protease (Mpro). HYPOTHESIS/PURPOSE: Following the finding that xanthohumol (XN) is a potent inhibitor of SARS-CoV-2 Mpro, the effect of XN and its major derivatives isoxanthohumol (IXN), 6-prenylnaringenin (6-PN), and 8-prenylnaringenin (8-PN) from hops on SARS-CoV-2 papain-like protease (PLpro) were investigated. STUDY DESIGN: The modulatory effect of the hop compounds on PLpro were studied first in silico and then in vitro. In addition, the actual effect of hop compounds on the replication of SARS-CoV-2 in host cells was investigated. METHODS: In silico docking analysis was used to predict the binding affinity of hop compounds to the active site of PLpro. A recombinant PLpro was cloned, purified, characterized, and analyzed by small-angle X-ray scattering (SAXS), deISGylation assays, and kinetic analyses. Antiviral activity of hop compounds was assessed using the fluorescently labeled wildtype SARS-CoV-2 (icSARS-CoV-2-mNG) in Caco-2 host cells. RESULTS: Our in silico docking suggests that the purified hop compounds bind to the active site of SARS-CoV-2 PLpro blocking the access of its natural substrates. The hop-derived compounds inhibit SARS-CoV-2 PLpro with half maximal inhibitory concentration (IC50) values in the range of 59-162 µM. Furthermore, we demonstrate that XN and 6-PN, in particular, impede viral replication with IC50 values of 3.3 µM and 7.3 µM, respectively. CONCLUSION: In addition to the already known inhibition of Mpro by XN, our results show, for the first time, that hop-derived compounds target also SARS-CoV-2 PLpro which is a promising therapeutic target as it contributes to both viral replication and modulation of the immune system. These findings support the possibility to develop new hop-derived antiviral drugs targeting human coronaviruses.

Bacterial OTU deubiquitinases regulate substrate ubiquitination upon Legionella infection.

Legionella pneumophila causes a severe pneumonia known as Legionnaires' disease. During the infection, Legionella injects more than 300 effector proteins into host cells. Among them are enzymes involved in altering the host-ubiquitination system. Here, we identified two LegionellaOTU (ovarian tumor)-like deubiquitinases (LOT-DUBs; LotB [Lpg1621/Ceg23] and LotC [Lpg2529]). The crystal structure of the LotC catalytic core (LotC14-310) was determined at 2.4 Å. Unlike the classical OTU-family, the LOT-family shows an extended helical lobe between the Cys-loop and the variable loop, which defines them as a unique class of OTU-DUBs. LotB has an additional ubiquitin-binding site (S1'), which enables the specific cleavage of Lys63-linked polyubiquitin chains. By contrast, LotC only contains the S1 site and cleaves different species of ubiquitin chains. MS analysis of LotB and LotC identified different categories of host-interacting proteins and substrates. Together, our results provide new structural insights into bacterial OTU-DUBs and indicate distinct roles in host-pathogen interactions.