muc1 recombinant proteins  (Novoprotein)

Journal: The Journal of Biological Chemistry

Article Title: The protease cathepsin K can debulk the cancer glycocalyx

doi: 10.1016/j.jbc.2026.111206

Figure Lengend Snippet: Only cathepsin K degrades cell-surface mucins on K562s. A, schematic describing the flow cytometry assay used to evaluate the degradation of cell-surface mucins on K562 cells by cathepsins. B, cell viability of cells following enzymatic treatment. Normalized staining for ( C ) MUC1, ( D ) CD43, and ( E ) total mucins via StcE E447D following enzymatic treatments for 3 h at pH 6 in Hanks' balanced salt solution (HBSS). Staining was normalized such that PBS-treated and fluorescence minus-one controls are defined as 100% and 0% staining within each replicate ( n = 3–4 biologically independent replicates). Cathepsins have been labeled with their letter, for example, “A” refers to cathepsin A. CTSE was excluded because of the low pH of the CTSE activation buffer causing cellular toxicity . See for representative flow cytometry histograms, assays performed at pH 5 and 7, and bar graphs with raw median fluorescence intensity (MFI) values. Data are shown as mean ± SD from three to four biologically independent replicates, and each dot represents a single replicate. p Values determined via one-way ANOVA corrected via Dunnett’s multiple comparison test, with a single pooled variance. ∗ p ≤ 0.05, ∗∗ p ≤ 0.01, ∗∗∗ ≤ 0.001, and ∗∗∗∗ p ≤ 0.0001. CTSE, cathepsin E; StcE E447D , inactive point mutant of StcE used as a pan-mucin probe.

Article Snippet: Recombinant PSGL-1 (10 μg; Sino Biological; 13863-H08H), recombinant MUC1 (Sino Biological; 12123-H05H), recombinant CD43 (R&D Systems; 9680-CD-050), fetuin (Promega; V4961), and purified MUC16 (purified from OVCAR-3 as above) were mixed with CTSK or PBS control at a 1:50 CTSK:substrate molar ratio in 1:1 PBS:100 mM sodium acetate with a final pH of 5 (buffer pHed as above).

Techniques: Flow Cytometry, Staining, Fluorescence, Labeling, Activation Assay, Comparison, Mutagenesis

Journal: The Journal of Biological Chemistry

Article Title: The protease cathepsin K can debulk the cancer glycocalyx

doi: 10.1016/j.jbc.2026.111206

Figure Lengend Snippet: CTSK degrades cell-surface mucins across multiple cell lines. A, schematic describing the flow cytometry assay for detecting the degradation of cell-surface mucins on H82, OVCAR3, MCF10A, and MCF10 MUC1 cells. B, median fluorescence intensity (MFI) of MUC1 and total mucins via StcE E447D . C, normalized MFI of MUC1 staining for MCF10 ± MUC1 following enzymatic treatments for 1 h a 37 °C at pH 6.75 in Hanks' balanced salt solution (HBSS). D, normalized StcE E447D staining for cell lines following enzymatic treatment for 1 h a 37 °C at pH 6.75. See , A – D for representative histograms. Data are shown as mean ± SD from two to four biologically independent replicates, and each dot represents a single replicate. P Values determined via ( C ) two-way ANOVA or ( D ) one-way ANOVA, both corrected via Tukey’s multiple comparison test, with a single pooled variance for each cell line. ∗ p ≤ 0.05, ∗∗ p ≤ 0.01, ∗∗∗ ≤ 0.001, and ∗∗∗∗ p ≤ 0.0001. CTSK, cathepsin K; MUC, mucin; StcE E447D , inactive point mutant of StcE used as a pan-mucin probe.

Article Snippet: Recombinant PSGL-1 (10 μg; Sino Biological; 13863-H08H), recombinant MUC1 (Sino Biological; 12123-H05H), recombinant CD43 (R&D Systems; 9680-CD-050), fetuin (Promega; V4961), and purified MUC16 (purified from OVCAR-3 as above) were mixed with CTSK or PBS control at a 1:50 CTSK:substrate molar ratio in 1:1 PBS:100 mM sodium acetate with a final pH of 5 (buffer pHed as above).

Techniques: Flow Cytometry, Fluorescence, Staining, Comparison, Mutagenesis

Journal: The Journal of Biological Chemistry

Article Title: The protease cathepsin K can debulk the cancer glycocalyx

doi: 10.1016/j.jbc.2026.111206

Figure Lengend Snippet: Cathepsin K (CTSK) tolerates glycans near the cleavage site. A, cleavage motif of CTSK was generated from mass spectrometry analysis of ( left ) glycopeptides, ( center ) nonmodified peptides, and ( right ) modified and nonmodified peptides generated from CTSK digestion of purified and recombinant mucins and nonmucin glycoproteins, followed by trypsin digestion (see the section). The bar graphs on top of the glycopeptide cleavage motif indicate the frequency of O -glycosylation at each threonine and serine residue at that position. B, top, visualization of CTSK cleavage sites in recombinant MUC1 residues 24–47. The sialylated core-1 glycan at specific resides indicates that glycans were seen at those sites. This specific glycan was seen often in the dataset, but its depiction here is only intended to indicate glycosites, not to represent the diversity of all glycans detected at each glycosite in the dataset. Purple diamond , sialic acid; yellow circle , galactose; yellow square , N-acetylgalactosamine; and yellow circle , glycosylation site. Colored bars represent individual detected peptide sequences from CTSK cleavage only, with any shared peptides with chymotrypsin removed. Bottom, annotated spectrum for the indicated MUC1 O -glycopeptide. C, top, visualization of CTSK cleavage sites in recombinant P-selectin glycoprotein ligand-1 (PSGL-1) residues 148 to 197, represented as in ( B ), but this time from the tryptic + CTSK dataset, with all tryptic cleavage sites removed. Bottom, annotated spectrum for the indicated PSGL-1 O -glycopeptide. MUC, mucin.

Article Snippet: Recombinant PSGL-1 (10 μg; Sino Biological; 13863-H08H), recombinant MUC1 (Sino Biological; 12123-H05H), recombinant CD43 (R&D Systems; 9680-CD-050), fetuin (Promega; V4961), and purified MUC16 (purified from OVCAR-3 as above) were mixed with CTSK or PBS control at a 1:50 CTSK:substrate molar ratio in 1:1 PBS:100 mM sodium acetate with a final pH of 5 (buffer pHed as above).

Techniques: Generated, Mass Spectrometry, Modification, Purification, Recombinant, Glycoproteomics, Residue

Journal: The Journal of Biological Chemistry

Article Title: The protease cathepsin K can debulk the cancer glycocalyx

doi: 10.1016/j.jbc.2026.111206

Figure Lengend Snippet: Cathepsin K (CTSK) sheds bulky glycan polymers across multiple cell lines. A, schematic describing the flow cytometry assay for detecting shedding of glycan polymers from H82, OVCAR3, MCF10A, and MCF10 MUC1 cells following enzymatic treatments with CTSK, heat-inactivated CTSK (HI CTSK), StcE, heparinase, chondroitinase, and the polySia-specific endosialidase (EndoNA). Cells were stained for ( B ) heparan sulfate using fibroblast growth factor 2 (FGF2), which is a probe for heparan sulfate, ( C ) polysialic acid using anti-polySia antibody (clone 735), ( D ) chondroitin sulfate using anti–chondroitin sulfate antibody (clone CS-56), and ( E ) viability following enzymatic treatment of cells for 1 h at 37 °C at pH 6.75. Staining was ( B ) normalized median fluorescence intensity (MFI) to 100% buffer control and 0% secondary only or ( C and D ) quantified as percent positive staining because of the broad and non-normal distribution of the cell populations. See for normalized staining values and representative histograms. F, change in glycocalyx thickness of YSCCC, MCF10A, and MCF10 MUC1 cells following enzymatic treatment relative to buffer control was measured using scanning angle interference microscopy. Each data point is the average of 20 to 50 individual cell measurements performed on a single day and represents an independent biological replicate. See for individual cell measurements. Data are shown as mean ± SD from two to five biologically independent replicates, and each dot represents a single replicate. p Values determined via one-way or two-way ANOVA corrected via Tukey’s multiple comparison test, with a single pooled variance for each cell line. ∗ p ≤ 0.05, ∗∗ p ≤ 0.01, ∗∗∗ ≤ 0.001, and ∗∗∗∗ p ≤ 0.0001.

Article Snippet: Recombinant PSGL-1 (10 μg; Sino Biological; 13863-H08H), recombinant MUC1 (Sino Biological; 12123-H05H), recombinant CD43 (R&D Systems; 9680-CD-050), fetuin (Promega; V4961), and purified MUC16 (purified from OVCAR-3 as above) were mixed with CTSK or PBS control at a 1:50 CTSK:substrate molar ratio in 1:1 PBS:100 mM sodium acetate with a final pH of 5 (buffer pHed as above).

Techniques: Glycoproteomics, Flow Cytometry, Staining, Fluorescence, Control, Microscopy, Comparison

Journal: ACS Omega

Article Title: Modified Sanliangsan Improved Sjogren’s Syndrome Complicated with Interstitial Lung Disease by Suppressing Serum MUC1 Levels

doi: 10.1021/acsomega.4c01147

Figure Lengend Snippet: SP treatment suppresses hub SS-related MUC1 expression to improve SS-ILD. (A, B) Volcano plots display the DEGs in the salivary gland (A) and parotid (B) of patients with SS. The hub genes associated with SS are labeled by a black arrow. (C) Venn diagram displays the hub DEGs shared in the salivary gland, peripheral blood, and parotid of patients with SS. The DEGs from the salivary gland, peripheral blood, and parotid are represented by orange, gray, and blue circles, respectively. (D) Topological protein–protein interaction network of hub DEGs associated with SS. (E) Relative expression levels of APP and MUC1 in patients with SS-ILD vs healthy individuals. (F) Expression level of MUC1 in serum of patients with SS-ILD before and after SP and CK treatments. (G) ELISA of the serum protein levels of MUC1 in patients with SS-ILD before and after SP and CK treatments; “NS: no significance; * P < 0.05; ** P < 0.01”.

Article Snippet: The recombinant MUC1 protein was obtained by GST-tag protein purification kit (Beyotime Biotechnology, Shanghai, China), then collected, and dialyzed in PBS glycerol buffer.

Techniques: Expressing, Labeling, Enzyme-linked Immunosorbent Assay

Journal: ACS Omega

Article Title: Modified Sanliangsan Improved Sjogren’s Syndrome Complicated with Interstitial Lung Disease by Suppressing Serum MUC1 Levels

doi: 10.1021/acsomega.4c01147

Figure Lengend Snippet: SP therapy improves SS-LID by affecting the regulation network of MUC1 . (A) Scatter plot displays the pathway enrichment results of transcriptome profiles of the peripheral blood of patients with SS. The plot size and color represent the gene number and significance of each pathway. (B) Protein–protein interaction network showing the potential regulator of MUC1. (C) Expression of genes interacting with MUC1 in the serum of patients with SS-ILD after SP therapy.

Article Snippet: The recombinant MUC1 protein was obtained by GST-tag protein purification kit (Beyotime Biotechnology, Shanghai, China), then collected, and dialyzed in PBS glycerol buffer.

Techniques: Expressing

Journal: ACS Omega

Article Title: Modified Sanliangsan Improved Sjogren’s Syndrome Complicated with Interstitial Lung Disease by Suppressing Serum MUC1 Levels

doi: 10.1021/acsomega.4c01147

Figure Lengend Snippet: Network pharmacology analysis of SP in improving SS-ILD. (A) Category of metabolites from SP. (B) KEGG pathways analysis on targets of bioactive compounds of SP. (C) Integrated network of bioactive compounds and related proteins. (D) Molecular docking analysis detects the interaction between MUC1 protein and eugenol. (E) Isothermal titration calorimetry (ITC) enthalpograms of eugenol binding to MUC1. Titration data are presented as blue plots and fit as a black solid line. (F) RT-qPCR determines the effects of eugenol in affecting abnormal upregulation of genes involved in SS-ILD pathogenesis.

Article Snippet: The recombinant MUC1 protein was obtained by GST-tag protein purification kit (Beyotime Biotechnology, Shanghai, China), then collected, and dialyzed in PBS glycerol buffer.

Techniques: Isothermal Titration Calorimetry, Binding Assay, Titration, Quantitative RT-PCR

Journal: STAR Protocols

Article Title: Protocol for characterizing the inhibition of SARS-CoV-2 infection by a protein of interest in cultured cells

doi: 10.1016/j.xpro.2022.101802

Figure Lengend Snippet: Representative bar graph of an experiment testing the effect of MUC1 on SARS-CoV-2 inhibition Mock means MUC1 is denatured by 100°C for 10 min. Data are represented as mean ± SEM. Data is reproduced from ( Lai et al., 2022 ).

Article Snippet: MUC1 , Novoprotein , Cat# CS58.

Techniques: Inhibition

Journal: STAR Protocols

Article Title: Protocol for characterizing the inhibition of SARS-CoV-2 infection by a protein of interest in cultured cells

doi: 10.1016/j.xpro.2022.101802

Figure Lengend Snippet: The inhibition of different stages of SARS-CoV-2 infection by MUC1 (A and B) Representative bar graph of an experiment testing the effects of MUC1 on SARS-CoV-2 attaching (A) and entry (B) to the cells. Data are represented as mean ± SEM. Data are reproduced from ( Lai et al., 2022 ).

Article Snippet: MUC1 , Novoprotein , Cat# CS58.

Techniques: Inhibition, Infection

Journal: STAR Protocols

Article Title: Protocol for characterizing the inhibition of SARS-CoV-2 infection by a protein of interest in cultured cells

doi: 10.1016/j.xpro.2022.101802

Figure Lengend Snippet: Representative bar graph of an experiment testing the effects of MUC1 and heparin of different combinations on SARS-CoV-2 attaching to the cell inhibition Data are represented as mean ± SEM. Data is reproduced from ( Lai et al., 2022 ).

Article Snippet: MUC1 , Novoprotein , Cat# CS58.

Techniques: Inhibition

Journal: STAR Protocols

Article Title: Protocol for characterizing the inhibition of SARS-CoV-2 infection by a protein of interest in cultured cells

doi: 10.1016/j.xpro.2022.101802

Figure Lengend Snippet:

Article Snippet: MUC1 , Novoprotein , Cat# CS58.

Techniques: Recombinant, Modification, Saline, Isolation, cDNA Synthesis, SYBR Green Assay, CCK-8 Assay, Software, Cell Culture, Sterility, Cell Counting, Real-time Polymerase Chain Reaction, Adhesive

Journal: STAR Protocols

Article Title: Protocol for characterizing the inhibition of SARS-CoV-2 infection by a protein of interest in cultured cells

doi: 10.1016/j.xpro.2022.101802

Figure Lengend Snippet:

Article Snippet: MUC1 , Novoprotein , Cat# CS58.

Techniques: Infection, Concentration Assay

Journal: STAR Protocols

Article Title: Protocol for characterizing the inhibition of SARS-CoV-2 infection by a protein of interest in cultured cells

doi: 10.1016/j.xpro.2022.101802

Figure Lengend Snippet:

Article Snippet: MUC1 , Novoprotein , Cat# CS58.

Techniques: