e. coli codon-optimized nucleotide sequence of smlt1473  (GenScript corporation)

Journal: Frontiers in Medicine

Article Title: Novel immune checkpoint inhibitor FilC/PD-1 recombinant vaccinia virus inhibits hepatocellular carcinoma

doi: 10.3389/fmed.2025.1622209

Figure Lengend Snippet: Structural representations of FilC and PD-1 proteins (a) FilC – Structural model of the FilC protein, depicting its folded β -sheet arrangement. (b) PD-1 – Structural model of the PD-1 immune checkpoint receptor.

Article Snippet: Codon-optimized nucleotide sequences for FilC (GenBank accession no. OR123456 ) and the PD-1 inhibitor (GenBank accession no. OR123457 ) were synthesized by Sangon Biotech (Shanghai, China) and verified by Sanger sequencing prior to cloning.

Techniques:

Journal: Frontiers in Medicine

Article Title: Novel immune checkpoint inhibitor FilC/PD-1 recombinant vaccinia virus inhibits hepatocellular carcinoma

doi: 10.3389/fmed.2025.1622209

Figure Lengend Snippet: Structural quality assessment of FilC and PD-1 Models (a) FilC Model Validation: Various structural validation metrics, including Rfree, Clashscore, Ramachandran outliers, Sidechain outliers, and RSRZ outliers, are shown. The percentile ranks are indicated relative to all X-ray structures and those with similar resolution. (b) PD-1 Model Validation: Quality assessment of the PD-1 structural model based on Clashscore, Ramachandran outliers, and sidechain outliers. The percentile ranks are provided relative to all NMR structures.

Article Snippet: Codon-optimized nucleotide sequences for FilC (GenBank accession no. OR123456 ) and the PD-1 inhibitor (GenBank accession no. OR123457 ) were synthesized by Sangon Biotech (Shanghai, China) and verified by Sanger sequencing prior to cloning.

Techniques: Biomarker Discovery

Journal: Frontiers in Medicine

Article Title: Novel immune checkpoint inhibitor FilC/PD-1 recombinant vaccinia virus inhibits hepatocellular carcinoma

doi: 10.3389/fmed.2025.1622209

Figure Lengend Snippet: Brightfield and fluorescence microscopy images showing vaccinia virus–mediated transgene expression. (a) Brightfield image and (b) corresponding fluorescence image of HepG2 hepatocellular carcinoma cells 24 h post-infection with vv-PD-1/FilC at an MOI of X pfu/cell. Red fluorescence indicates reporter expression (e.g., mCherry) under the control of the viral promoter. Scale bars: 50 μm. (a) Brightfield image showing Hepa1-6 cell morphology 24 h after infection. (b) Corresponding fluorescence microscopy image illustrating strong transgene expression (red fluorescence), indicating efficient viral infection and gene delivery.

Article Snippet: Codon-optimized nucleotide sequences for FilC (GenBank accession no. OR123456 ) and the PD-1 inhibitor (GenBank accession no. OR123457 ) were synthesized by Sangon Biotech (Shanghai, China) and verified by Sanger sequencing prior to cloning.

Techniques: Fluorescence, Microscopy, Virus, Expressing, Infection, Control

Journal: Frontiers in Medicine

Article Title: Novel immune checkpoint inhibitor FilC/PD-1 recombinant vaccinia virus inhibits hepatocellular carcinoma

doi: 10.3389/fmed.2025.1622209

Figure Lengend Snippet: Time-dependent viral replication kinetics of the FilC/PD-1 recombinant vaccinia virus in HCC and non-cancerous cell lines. Statistical analysis was performed using one-way ANOVA with Tukey’s post hoc test; error bars represent SD ( n = 3); p < 0.01 compared to non-HCC cell lines at 48 h and 72 h.

Article Snippet: Codon-optimized nucleotide sequences for FilC (GenBank accession no. OR123456 ) and the PD-1 inhibitor (GenBank accession no. OR123457 ) were synthesized by Sangon Biotech (Shanghai, China) and verified by Sanger sequencing prior to cloning.

Techniques: Recombinant, Virus

Journal: Frontiers in Medicine

Article Title: Novel immune checkpoint inhibitor FilC/PD-1 recombinant vaccinia virus inhibits hepatocellular carcinoma

doi: 10.3389/fmed.2025.1622209

Figure Lengend Snippet: Differential gene expression between FilC/PD-1–treated tumors and controls.

Article Snippet: Codon-optimized nucleotide sequences for FilC (GenBank accession no. OR123456 ) and the PD-1 inhibitor (GenBank accession no. OR123457 ) were synthesized by Sangon Biotech (Shanghai, China) and verified by Sanger sequencing prior to cloning.

Techniques: Gene Expression

Journal: Nature Communications

Article Title: Ultrapotent SARS coronavirus-neutralizing single-domain antibodies that clamp the spike at its base

doi: 10.1038/s41467-025-60250-1

Figure Lengend Snippet: a left: model of full-length prefusion S (6VSB_1_1_2 ) with superimposition of the R3DC23 – HR2 complex, all shown in molecular surface representation, with N-glycans in stick representation. R3DC23 VHHs are colored in orange, labeled S protein regions: cytoplasmic domain (red; CP), transmembrane domain (gray; TM), heptad repeat 2 (blue; HR2), S2 stem helix (deep pink; SH), heptad repeat 1 (lemon, HR1), central helix and connector domain (pink; CH-CD), the S1 regions encompassing the N-terminal domain (NTD) and receptor binding domain (RBD) (light blue). Right: model of the proteolytically processed postfusion S2 subunit (7E9T ), color coded as in prefusion spike. b Side and axial view (inset) of the R3DC23 – HR2 complex (sand and blue, respectively) superimposed with prefusion HR2 coiled-coil (light blue). In R3DC23, CDR1, 2, and 3 are colored magenta, yellow, and cyan, respectively. The HR2-binding epitope spanning D1192 – Y1206 is shown in stick representation. c Close-up of boxed region, encompassing a single VHH and two HR2 copies (i and ii) forming the adjoined binding epitope. Escape mutant positions are labeled in red. d Axial view of HR1-HR2 (lemon and blue) region of postfusion S protein, superimposed with R3DC23 in the HR2 complex (gray). e The amino acids involved in interactions between R3DC23 and two HR2 helices as observed in the crystal of the R3DC23-HR2 complex are indicated in red.

Article Snippet: For the pCG1-SARS-2-BA.2 Sdel18 expression vector, a codon-optimized spike protein nucleotide sequence containing the BA.2 mutations (T19I, ΔL14-P26, A27S, G142D, V213G, G339D, S371F, S373P, S375F, T376A, D405N, R408S, K417N, N440K, S477N, T478K, E484A, Q493R, Q498R, N501Y, Y505H, D614G, H655Y, N679K, P681H, N764K, D796Y, Q954H, N969K) and flanking BamH I and Sal I restriction sites was ordered at GeneArt (Thermo Fischer Scientific) and cloned in the pCG1 vector as a Bam HI/ Sal I fragment.

Techniques: Labeling, Binding Assay, Mutagenesis

Journal: Applied and Environmental Microbiology

Article Title: Applying a polysaccharide lyase from Stenotrophomonas maltophilia to disrupt alginate exopolysaccharide produced by Pseudomonas aeruginosa clinical isolates

doi: 10.1128/aem.01853-24

Figure Lengend Snippet: Smlt1473 inhibits the mucoid phenotype of P. aeruginosa , but data suggest most effective enzyme concentration varies by isolate. ( A ) Representative image of phenotypic changes for P. aeruginosa UVA 44618 in the presence and absence of Smlt1473 indicating an inhibitory function. ( B–F ) Each isolate of P. aeruginosa was grown in the presence and absence of Smlt1473, plate contents were collected, and uronic acid concentration, which corresponds to alginate content, was quantified. Uronic acid concentration was determined by measuring the absorbance at 530 nm (A530) of the resulting solution. High A530 corresponds to high alginate content, whereas low A530 corresponds to low alginate content. Y222F is the catalytically inactive form of Smlt1473 used to show that results are due to an active enzyme. The results are means and standard deviations and statistical analysis was performed using a One-way Welch’s ANOVA and Dunnett’s T3 multiple comparison post-hoc test.

Article Snippet: An E. coli codon-optimized nucleotide sequence of Smlt1473 (GenBank accession number CAQ45011 ) was synthesized and cloned into pET-28a(+) using NcoI/XhoI restriction sites (GenScript).

Techniques: Concentration Assay, Comparison

Journal: Applied and Environmental Microbiology

Article Title: Applying a polysaccharide lyase from Stenotrophomonas maltophilia to disrupt alginate exopolysaccharide produced by Pseudomonas aeruginosa clinical isolates

doi: 10.1128/aem.01853-24

Figure Lengend Snippet: SEM images of all mucoid P. aeruginosa isolates treated with Smlt1473 and buffer showing enzymatic inhibition of mucoid phenotype. Samples were grown in the presence of enzyme or buffer, transferred to 12 mm glass slides, glutaraldehyde fixed, and imaged using SEM. Each set of images is shown at 50,000× magnification. The left side of the figure panel depicts samples that were grown in the presence of buffer, whereas the right side shows samples that were grown in the presence of Smlt1473. (A and B) UVA 44618, (C and D) UVA 61605, (E and F) UVA 84977, (G and H) UVA 55009, and (I and J) PDO300.

Article Snippet: An E. coli codon-optimized nucleotide sequence of Smlt1473 (GenBank accession number CAQ45011 ) was synthesized and cloned into pET-28a(+) using NcoI/XhoI restriction sites (GenScript).

Techniques: Inhibition

Journal: Applied and Environmental Microbiology

Article Title: Applying a polysaccharide lyase from Stenotrophomonas maltophilia to disrupt alginate exopolysaccharide produced by Pseudomonas aeruginosa clinical isolates

doi: 10.1128/aem.01853-24

Figure Lengend Snippet: Smlt1473 degrades the mucoid biofilm of P. aeruginosa after it has been established on a surface. ( A ) UVA 61605 biofilm after 24 h of growth showing a prominent raised, mucoid phenotype. ( B–F ) Each P. aeruginosa isolate was grown on LB agar for 24 h at 37°C with no treatment to develop an established mucoid phenotype, as shown in panel A. Plate contents were collected, and the alginate-containing biofilm was used as the substrate in the TBA assay. Upon addition of enzyme, alginate is depolymerized via β-elimination mechanism where unsaturated products react with thiobarbituric acid to create a pink chromogen with absorbance at 540 nm. High A540 corresponds to greater alginate depolymerization, and low A540 corresponds to minimal alginate depolymerization. The results presented are means and standard deviations, and statistical analysis was performed using a one-way Welch’s ANOVA and Dunnett’s T3 multiple comparison post-hoc tests. ( G ) Representative image displaying the pink chromogen as a result of the addition of Smlt1473 to UVA 55009 biofilm mixture (right) compared with the addition of buffer to the mixture (left).

Article Snippet: An E. coli codon-optimized nucleotide sequence of Smlt1473 (GenBank accession number CAQ45011 ) was synthesized and cloned into pET-28a(+) using NcoI/XhoI restriction sites (GenScript).

Techniques: Comparison

Journal: Applied and Environmental Microbiology

Article Title: Applying a polysaccharide lyase from Stenotrophomonas maltophilia to disrupt alginate exopolysaccharide produced by Pseudomonas aeruginosa clinical isolates

doi: 10.1128/aem.01853-24

Figure Lengend Snippet: Stacked 1 H NMR spectra of all five P. aeruginosa isolates showing acetylation and quantitative determination of degree of acetylation. ( A ) Peaks around 2.12ppm are a result of the acetyl group of acetylated sugars, indicating all of the isolates in this study are comprised of acetylated alginate. ( B ) The degree of acetylation was quantitatively determined using a method previously described, further proving that all biofilm samples have some fraction of acetylated alginate. Data in shows that Smlt1473 degrades established mucoid biofilm, and in combination with these NMR and degree of acetylation results, we can conclude that Smlt1473 is effective against acetylated alginate.

Article Snippet: An E. coli codon-optimized nucleotide sequence of Smlt1473 (GenBank accession number CAQ45011 ) was synthesized and cloned into pET-28a(+) using NcoI/XhoI restriction sites (GenScript).

Techniques: