Crystal structure and activity profiling of deubiquitinating inhibitors-bound to SARS-CoV-2 papain like protease revealed new allosteric sites for antiviral therapies

Emerging variants of SARS-CoV-2 still threaten the effectiveness of currently deployed vaccines, and antivirals can prove to be an effective therapeutic option for attenuating it. The papain-like protease (PLpro) is an attractive target due to its sequence conservation and critical role in the replication and pathogenesis of SARS-CoV-2. PLpro also plays very important role in modulation of host immune responses by deubiquitinating (DUBs) or deISGylating host proteins. Thus, targeting PLpro serves as a two-pronged approach to abate SARS-CoV-2. Due to its structural and functional similarities with the host DUB enzymes, an in-house library of DUB inhibitors was constituted in this study. Five promising compounds exhibiting high binding affinities with the substrate binding site of PLpro were identified from a library of 81 compounds with in silico screening, docking, and simulation studies. Interestingly, lithocholic acid, linagliptin, teneligliptin, and flupenthixol significantly inhibited the proteolytic activity of PLpro. Each of these compounds abrogated in vitro replication of SARS-CoV-2 with EC50 values in the range of 5-21 M. In addition, crystal structure of SARS-CoV-2 PLpro and its complex with inhibitors have been determined that revealed their inhibitory mechanism. The findings of this study provide the proof-of-principle that the DUB inhibitors hold high potential as a new class of therapeutics against SARS-CoV-2. Additionally, this is the first study that has opened a new avenue towards not only targeting PLpro active site but also simultaneously directing towards restoration of antiviral immune response of the host for deterring SARS-CoV-2. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=173 SRC="FIGDIR/small/516107v1_ufig1.gif" ALT="Figure 1"> View larger version (50K): org.highwire.dtl.DTLVardef@90c3fborg.highwire.dtl.DTLVardef@32b4daorg.highwire.dtl.DTLVardef@ff858dorg.highwire.dtl.DTLVardef@198fce7_HPS_FORMAT_FIGEXP M_FIG C_FIG

An In vitro Comparison and Evaluation of Sealing Ability of Newly Introduced C-point System, Cold Lateral Condensation, and Thermoplasticized Gutta-Percha Obturating Technique: A Dye Extraction Study.

AIM: The aim of this study is to compare and to evaluate sealing ability of newly introduced C-point system, cold lateral condensation, and thermoplasticized gutta-percha obturating technique using a dye extraction method. MATERIALS AND METHODOLOGY: Sixty extracted maxillary central incisors were decoronated below the cementoenamel junction. Working length was established, and biomechanical preparation was done using K3 rotary files with standard irrigation protocol. Teeth were divided into three groups according to the obturation protocol; Group I-Cold lateral condensation, Group II-Thermoplasticized gutta-percha, and Group III-C-Point obturating system. After obturation all samples were subjected to microleakage assessment using dye extraction method. Obtained scores will be statistical analyzed using ANOVA test and post hoc Tukey's test. RESULTS: One-way analysis of variance revealed that there is significant difference among the three groups with P value (0.000 < 0.05). Tukey's HSD post hoc tests for multiple comparisons test shows that the Group II and III perform significantly better than Group I. Group III performs better than Group II with no significant difference. CONCLUSION: All the obturating technique showed some degree of microleakage. Root canals filled with C-point system showed least microleakage followed by thermoplasticized obturating technique with no significant difference among them. C-point obturation system could be an alternative to the cold lateral condensation technique.

Investigate the correlation between clinical sign and symptoms and the presence of P. gingivalis, T. denticola, and T. forsythia individually or as a "Red complex" by a multiplex PCR method.

AIM: The aim of this study was to investigate the correlation between endodontic clinical signs and symptoms and the presence of Porphyromonas gingivalis, Treponema denticola, and Tannerella forsythia or their association by Multiplex polymerase chain reaction assay. MATERIALS AND METHODS: Microbial samples were taken from 30 cases with necrotic pulp tissues in primary infections. DNA was extracted from the samples, which were analyzed for the presence of three endodontic pathogens by using species-specific primers. RESULTS: P. gingivalis, T. denticola, T. forsythia, and Red Complex were present in 11, 17, 4, and 2 canals, respectively. Clinical and statistically significant relationships were found between T. forsythia and mobility and between T. denticola and swelling. (P < 0.05). Presence of other Red complex bacteria shows clinical association with presence of signs and symptoms but no statistically significant relationship. CONCLUSION: The high prevalence of P. gingivalis, T. denticola, and T. forsythia in the examined samples suggests that these bacteria are related to the etiology of symptomatic periradicular diseases.