Journal: Journal of Medicinal Chemistry

Article Title: Design of a Fluorescence Polarization Probe for Enterovirus 2C Proteins

doi: 10.1021/acs.jmedchem.5c01219

Figure Lengend Snippet: Genome structure of enterovirus, structure of 2C, and reported 2C inhibitors. (A) The EV genome contains ORF encoding structural proteins VP1, VP2, VP3, and VP4 and nonstructural proteins 2A, 2B, and 2C and 3A, 3B, 3C, and 3D. (B) Alignment of the crystal structure of CVB3 2C in complex with SFX (PDB ID 6S3A , colored in gray) and EV-A71 2C in complex with ATPγS (PDB ID 5GRB , colored in wheat). The C-terminal part of 2C is highlighted in orange, the zinc finger domain is highlighted in green, ATPγS is shown in blue sticks, and SFX is shown in yellow sticks. (C) Chemical structures of reported enterovirus 2C inhibitors.

Article Snippet: The enterovirus strains used in this study were obtained from commercial sources: EV-D68 US/MO/14-18947 (ATCC, NR-49129), EV-A71 Tainan/4643/1998 (BEI Resources, NR-471), and CVB3 strain Nancy (ATCC, VR-30).

Techniques:

Journal: Journal of Medicinal Chemistry

Article Title: Design of a Fluorescence Polarization Probe for Enterovirus 2C Proteins

doi: 10.1021/acs.jmedchem.5c01219

Figure Lengend Snippet: Design of the 2C FP probe. (A) FP assay principle. (B) SAR study and design of the 2C FP probe Jun14157 . (C) Superimposed structures of EV-D68 2C (homology model) with CVB3 2C in complex with SFX (PDB ID 6T3W ). EV-D68 2C is colored in green, CVB3 2C is colored in magenta, and the cavity of the allosteric pocket is colored in cyan. SFX is shown as wheat sticks. (D) The superposition of docking poses of Jun571 , Jun1377 , and Jun14157 occurred at the allosteric site of EV-D68 2C. 2C is colored in split pea, Jun571 is shown as gray sticks, Jun1377 is shown as marine sticks, and Jun14157 is shown as pink sticks.

Article Snippet: The enterovirus strains used in this study were obtained from commercial sources: EV-D68 US/MO/14-18947 (ATCC, NR-49129), EV-A71 Tainan/4643/1998 (BEI Resources, NR-471), and CVB3 strain Nancy (ATCC, VR-30).

Techniques: FP Assay

Journal: Journal of Medicinal Chemistry

Article Title: Design of a Fluorescence Polarization Probe for Enterovirus 2C Proteins

doi: 10.1021/acs.jmedchem.5c01219

Figure Lengend Snippet: Optimization of the FP assay using the Jun14157 probe. (A) Binding curves of the Jun14157 probe at 20, 50, and 100 nM with increasing concentrations of EV-D68 2C after 30 min of incubation. K d values represent means ± standard deviation (SD) from triplicate experiments. (B) Effect of dimethyl sulfoxide (DMSO) concentration on the mP values of bound and free probes. The assay was performed in triplicate. (C) Binding curves of 50 nM Jun14157 with increasing concentrations of WT and EV-D68 2C mutants D183V, D323G, and D183V/D323G. K d values are the mean ± SD from duplicate experiments. (D–F) FP titration curves of Jun14157 with EV-D68 2C-WT (D), EV-A71 2C (E), and CVB3 2C (F) with different incubation times. K d values are presented as the mean ± SD from triplicate experiments. Each data point represents the mean ± SD.

Article Snippet: The enterovirus strains used in this study were obtained from commercial sources: EV-D68 US/MO/14-18947 (ATCC, NR-49129), EV-A71 Tainan/4643/1998 (BEI Resources, NR-471), and CVB3 strain Nancy (ATCC, VR-30).

Techniques: FP Assay, Binding Assay, Incubation, Standard Deviation, Concentration Assay, Titration