Human infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes coronavirus disease 2019 (COVID-19) and there is no cure currently. The 3CL protease (3CLpro) is a highly conserved protease which is indispensable for CoVs replication, and is a promising target for development of broad-spectrum antiviral drugs. In this study we investigated the anti-SARS-CoV-2 potential of Shuanghuanglian preparation, a Chinese traditional patent medicine with a long history for treating respiratory tract infection in China. We showed that either the oral liquid of Shuanghuanglian, the lyophilized powder of Shuanghuanglian for injection or their bioactive components dose-dependently inhibited SARS-CoV-2 3CLpro as well as the replication of SARS-CoV-2 in Vero E6 cells. Baicalin and baicalein, two ingredients of Shuanghuanglian, were characterized as the first noncovalent, nonpeptidomimetic inhibitors of SARS-CoV-2 3CLpro and exhibited potent antiviral activities in a cell-based system. Remarkably, the binding mode of baicalein with SARS-CoV-2 3CLpro determined by X-ray protein crystallography was distinctly different from those of known 3CLpro inhibitors. Baicalein was productively ensconced in the core of the substrate-binding pocket by interacting with two catalytic residues, the crucial S1/S2 subsites and the oxyanion loop, acting as a “shield” in front of the catalytic dyad to effectively prevent substrate access to the catalytic dyad within the active site. Overall, this study provides an example for exploring the in vitro potency of Chinese traditional patent medicines and effectively identifying bioactive ingredients toward a specific target, and gains evidence supporting the in vivo studies of Shuanghuanglian oral liquid as well as two natural products for COVID-19 treatment.

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(-)-Stepholidine (l-SPD), an active ingredient of the Chinese herb Stephania, is the first compound found to have a dual function as a dopamine receptor D1 agonist and D2 antagonist. The preliminary dynamical behaviors of D1R and D2R and their interaction modes with l-SPD were investigated in our previous study. Recently, the pharmacological effect of l-SPD on D3R was elucidated as an antagonist. This new discovery in combination with the explosion of structural biology in GPCR superfamily prompted us to perform a more comprehensive investigation on the special pharmacological profiles of l-SPD on dopamine receptors. In this study, the integration of homology modeling, automated molecular docking, and MD simulations was used to probe the agonistic and antagonistic mechanism of l-SPD on D1R, D2R, and D3R. Our analyses showed that hydrogen bonding of the hydroxyl group on the D ring of l-SPD with side chain of N6.55 which, in combination with hydrophobic stacking between I3.40, F6.44 and W6.48, is the key feature to mediate the agonist effect of l-SPD on D1R, whereas the absence of hydrophobic stacking between I3.40, F6.44, and W6.48 in D2R and D3R excludes receptor activation. Finally, the agonistic and antagonistic mechanisms of l-SPD and an activation model of D1R were proposed on the basis of these findings. The present study could guide future experimental works on these receptors and has the significance to the design of functionally selective drugs targeting dopamine receptors.

The aspartate kinase (AK) from Mycobacterium tuberculosis (Mtb) catalyzes the biosynthesis of aspartate family amino acids, including lysine, threonine, isoleucine and methionine. We determined the crystal structures of the regulatory subunit of aspartate kinase from Mtb alone (referred to as MtbAKβ) and in complex with threonine (referred to as MtbAKβ-Thr) at resolutions of 2.6 Å and 2.0 Å, respectively. MtbAKβ is composed of two perpendicular non-equivalent ACT domains [aspartate kinase, chorismate mutase, and TyrA (prephenate dehydrogenase)] per monomer. Each ACT domain contains two α helices and four antiparallel β strands. The structure of MtbAKβ shares high similarity with the regulatory subunit of the aspartate kinase from Corynebacterium glutamicum (referred to as CgAKβ), suggesting similar regulatory mechanisms. Biochemical assays in our study showed that MtbAK is inhibited by threonine. Based on crystal structure analysis, we discuss the regulatory mechanism of MtbAK.

Avian infectious bronchitis virus (IBV) is a member of the group III coronaviruses, which differ from the other groups of coronaviruses in that they do not encode the essential pathogenic factor nonstructural protein 1 (nsp1) and instead start with nsp2. IBV nsp2 is one of the first replicase proteins to be translated and processed in the viral life cycle; however, it has an entirely unknown function. In order to better understand the structural details and functional mechanism of IBV nsp2, the recombinant protein was cloned, overexpressed in Escherichia coli, purified and crystallized. The crystals diffracted to 2.8 Å resolution and belonged to space group P2 1 , with unit-cell parameters a = 57. 0, b = 192.3, c = 105.7 Å , = 90.8 . Two molecules were found in the asymmetric unit; the Matthews coefficient was 3.9 Å 3 Da À1, corresponding to a solvent content of 68.2%.

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