The relationship between the conformation of interferon-α (IFN-α) and its anti-viral activity were analyzed by circular dichroism (CD) and flow cytometry (FCM) techniques. The recombinant human IFN-α (rIFN-α2b and rIFN-α2a) were used. CD spectra from 190 nm to 240 nm indicated that two the IFN-α showed stable secondary structure at 65 degrees C, but unstable when the temperature was above 65 degrees C, and the change was irreversible. FCM data of the anti-viral activity of IFN-α indicated that the change of its secondary structures partly weakened its anti-viral activity. The rIFN-α2b and rIFN-α2a showed the same phenomenon. These data indicated that the conformation of IFN-α is one of the factors to influence its anti-viral activity and the combination of CD and FCM is a good method to analyze the relationship between the conformation of protein drugs and their biological activities in single cell level.
Antiviral Agents ; chemistry ; Circular Dichroism ; Flow Cytometry ; Humans ; Interferon-alpha ; chemistry ; Protein Structure, Secondary ; Recombinant Proteins ; chemistry

Objective:To analyze the diagnostic and prognostic value of heparin-binding protein (HBP) combined with procalcitonin (PCT) and high sensitivity C-reactive protein (hs-CRP) in early sepsis.Methods:The clinical data of 116 patients with sepsis admitted to Zhejiang Xin′an International Hospital from August 2018 to December 2019 were retrospectively analyzed. They were divided into mild sepsis group (40 cases), severe sepsis group (46 cases) and septic shock group (30 cases) according to the severity of illness. According to the clinical prognosis, they were divided into survival group (87 cases) and death group (29 cases). In the same period, 104 outpatients were selected as control group. The levels of HBP, PCT and hs-CRP were compared in each group and the guiding value of HBP, PCT and hs-CRP in the diagnosis of early sepsis were analyzed by receiver operating characteristic (ROC) curve.Results:The levels of HBP, PCT and hs-CRP in sepsis group were higher than those in control group: (83.45 ± 11.29) μg/L vs. (4.81 ± 0.62) μg/L, (8.19 ± 1.14) μg/L vs. (0.24 ± 0.03) μg/L, (87.94 ± 13.58) mg/L vs. (1.34 ± 0.15) mg/L, and the differences were statistically significant ( P<0.05). The levels of HBP, PCT and hs-CRP were gradually increased with the aggravation of disease, and the differences were statistically significant ( P<0.05). The levels of HBP, PCT and hs-CRP in survival group were lower than those in death group: (67.94 ± 8.91) μg/L vs. (129.98 ± 18.43) μg/L, (6.46 ± 0.93) μg/L vs. (13.38 ± 1.77) μg/L, (65.19 ± 10.62) mg/L vs. (129.19 ± 22.46) mg/L, and the differences were statistically significant ( P<0.05). The ROC curve analysis showed that the area under the curve of HBP, PCT, hs-CRP and HBP + PCT + hs-CRP were 0.828, 0.835, 0.787 and 0.940, the sensitivity was 72.4%, 69.0%, 79.3% and 86.2%, and the specificity was 85.1%, 88.5%, 74.7% and 90.8%. Conclusions:HBP, PCT and hs-CRP joint detection have certain guiding value in the diagnosis of early sepsis, and can reflect the disease severity and prognosis.

ObjectiveBased on the experience of traditional quality evaluation， the quality of Atractylodis Macrocephalae Rhizoma（AMR） with different production methods such as direct seeding， transplanting after seedling raising， topping and non-topping， and difference in growth years was compared. MethodVernier caliper was used to measure the trait data of AMR in different production methods. Paraffin sections of AMR with different production methods were made by saffron solid green staining， and the microstructure was observed. The contents of water-soluble and alcohol-soluble extracts in AMR with different production methods were determined according to the 2020 edition of Chinese Pharmacopoeia. The content of water-soluble total polysaccharides in AMR with different production methods was detected by sulfuric acid-anthrone method. Fiber analyzer was used to detect the content of fiber components in AMR with different production methods. The contents of monosaccharides， oligosaccharides and some secondary metabolites in AMR with different production methods were detected by ultra performance liquid chromatography（UPLC）， and the differences of chemical components were compared by multivariate statistical analysis methods such as principal component analysis（PCA） and partial least squares-discriminant analysis（PLS-DA）. ResultIn terms of traits， the 3-year-old AMR with direct seeding and without topping was close to the high-quality AMR with "phoenix-head and crane-neck， strong sweetness and clear aroma" recorded in ancient materia medica， followed by the 3-year-old AMR with topping after transplanting， while the 2-year-old AMR with topping after transplanting with high market circulation rate was generally fat and strong with mild odor. In the microscopic aspect， the arrangement of xylem vessels and fiber bundles in the 3-year-old samples formed two obvious rings. Compared with the 2-year-old samples cultivated in Bozhou and Zhejiang， the 3-year-old samples without topping after transplanting had more wood fibers. In terms of chemical composition， the contents of 70% ethanol extract， fructose， glucose， sucrose， 1-kestose， atractylenolide Ⅰ， chlorogenic acid， neochlorogenic acid， cryptochlorogenic acid and other components in 3-year-old AMR with direct seeding and without topping were significantly higher than those in the other three samples（P<0.05）. The contents of cellulose， 70% ethanol extract， sucrose， atractylenolide Ⅰ， atractylone and other components in 3-year-old AMR with topping after transplanting were significantly higher than those in the 2-year-old AMR with high market circulation rate（P<0.05）， while the contents of water-soluble extract and water-soluble total polysaccharides in 2-year-old samples with topping after transplanting were significantly higher than those in the 3-year-old AMR with topping after transplanting， direct seeding and without topping（P<0.05）. ConclusionUnder the current mainstream production mode， too much manual intervention makes AMR heavily enriched in polysaccharides and increased the yield， but the accumulation of sweet substances， fragrant substances and fiber substances is insufficient， which affects its quality. The current quality standard of AMR has some shortcomings in guiding the high quality production of it， it is suggested to revise the quality standard of AMR， supplement the quantitative analysis of secondary metabolites， and strengthen the production of imitation wild AMR.

The global COVID-19 coronavirus pandemic has infected over 109 million people, leading to over 2 million deaths up to date and still lacking of effective drugs for patient treatment. Here, we screened about 1.8 million small molecules against the main protease (M^pro) and papain like protease (PL^pro), two major proteases in severe acute respiratory syndrome-coronavirus 2 genome, and identified 1851M^pro inhibitors and 205 PL^pro inhibitors with low nmol/l activity of the best hits. Among these inhibitors, eight small molecules showed dual inhibition effects on both M^pro and PL^pro, exhibiting potential as better candidates for COVID-19 treatment. The best inhibitors of each protease were tested in antiviral assay, with over 40% of M^pro inhibitors and over 20% of PL^pro inhibitors showing high potency in viral inhibition with low cytotoxicity. The X-ray crystal structure of SARS-CoV-2 M^pro in complex with its potent inhibitor 4a was determined at 1.8 Å resolution. Together with docking assays, our results provide a comprehensive resource for future research on anti-SARS-CoV-2 drug development.
Humans ; Antiviral Agents/chemistry* ; COVID-19 ; COVID-19 Drug Treatment ; High-Throughput Screening Assays ; Molecular Docking Simulation ; Protease Inhibitors/chemistry* ; SARS-CoV-2/enzymology* ; Viral Nonstructural Proteins