Journal: Frontiers in Oncology

Article Title: OTUB2 Facilitates Tumorigenesis of Gastric Cancer Through Promoting KDM1A-Mediated Stem Cell-Like Properties

doi: 10.3389/fonc.2021.711735

Figure Lengend Snippet: Otubain 2 (OTUB2) increases the stability of KDM1A. (A) Relative protein levels of KDM1A in GC cells transfected with OTUB2 overexpression plasmids (MKN-45 and SGC-7901 cells) or shRNA targeting OTUB2 (HGC-27 and AGS cells). Actin served as internal control. (B) co-immunoprecipitation was conducted with anti-Flag (Flag-OTUB2) or anti-HA (HA-KDM1A) antibody and was analyzed by western blotting in 293T cells. (C) after transfected with shOTUB2, cells were treated with or without MG132. The ubiquitination level of KDM1A in MKN-45 and HGC-27 cells was measured by Western blot. Actin served as internal control. (D) After transfected with OTUB2 overexpression plasmids (MKN-45 cells) or shOTUB2, cells were treated with or without cycloheximide (CHX) for 0, 1, 2, 4, and 8 h, respectively. The protein expression levels of KDM1A were assessed by Western blot. Actin was used as internal control. Data was represented as mean ± SD. * P < 0.05, ** P < 0.01, and *** P < 0.001.

Article Snippet: For deubiquitination assay, polyUb was purified by IP using anti-KDM1A antibody (1 μg, Boster, Catalog No. M00532; Beijing, China) from cells transfected with OTUB2 shRNA after treated with MG-132.

Techniques: Transfection, Over Expression, shRNA, Control, Immunoprecipitation, Western Blot, Ubiquitin Proteomics, Expressing

Journal: Frontiers in Oncology

Article Title: OTUB2 Facilitates Tumorigenesis of Gastric Cancer Through Promoting KDM1A-Mediated Stem Cell-Like Properties

doi: 10.3389/fonc.2021.711735

Figure Lengend Snippet: The expression of otubain 2 (OTUB2) and KDM1A in GC cells are positively correlated. (A) The expression of OTUB2, KDM1A, and CD44 was measured by immunohistochemistry in GC tissues collected from patients with GC. (B) The correlation between the expression of OTUB2, KDM1A, and CD44 in human tumor tissues was analyzed. (C) After OTUB2 was overexpressed (SGC-7901 cells) or downregulated (HGC-27 cells), the expression of KDM1A and CD44 was detected by immunohistochemistry in tumor tissues obtained from nude mice.

Article Snippet: For deubiquitination assay, polyUb was purified by IP using anti-KDM1A antibody (1 μg, Boster, Catalog No. M00532; Beijing, China) from cells transfected with OTUB2 shRNA after treated with MG-132.

Techniques: Expressing, Immunohistochemistry

Journal: Frontiers in Oncology

Article Title: OTUB2 Facilitates Tumorigenesis of Gastric Cancer Through Promoting KDM1A-Mediated Stem Cell-Like Properties

doi: 10.3389/fonc.2021.711735

Figure Lengend Snippet: OTUB2 promotes GC stem cell properties through regulating KDM1A expression. (B) After MKN-45 cells were co-transfected with OTUB2 overexpression plasmids/NC and shKDM1A/Scr. The HGC-27 cells were co-transfected with shOTUB2/shNC and OV-KDM1A/vector, the cell viability was detected by MTT assay (A) ; The spheroid-forming ability of GC cells were detected by sphere formation assay (B) . Wound healing assays (C) and Transwell assays (D) were used to determine the changes in migratory and invasive abilities of GC cells; Relative protein levels of KDM1A, Nanog, ALDH, and Oct4 IN GC cells were measured by Western blot (E) . Data was represented as mean ± SD. * P < 0.05, ** P < 0.01, and *** P < 0.001. # P < 0.05, ## P < 0.01, and ### P < 0.001.

Article Snippet: For deubiquitination assay, polyUb was purified by IP using anti-KDM1A antibody (1 μg, Boster, Catalog No. M00532; Beijing, China) from cells transfected with OTUB2 shRNA after treated with MG-132.

Techniques: Expressing, Transfection, Over Expression, Plasmid Preparation, MTT Assay, Tube Formation Assay, Western Blot

Journal: BMC Bioinformatics

Article Title: Investigating heterogeneous protein annotations toward cross-corpora utilization

doi: 10.1186/1471-2105-10-403

Figure Lengend Snippet: Experimental results of the GENIA subcategory annotations plus AIMed and GENETAG, respectively

Article Snippet: 4 , GENETAG+ DNA , Exact , 69.82 , 36.44 , 47.89.

Techniques:

Journal: Theoretical Biology & Medical Modelling

Article Title: Protein-lipid interactions: correlation of a predictive algorithm for lipid-binding sites with three-dimensional structural data

doi: 10.1186/1742-4682-3-17

Figure Lengend Snippet: Characteristics of the three-dimensional structures. Coordinate files were obtained from the Protein Data Bank [20]; 1HCI [28]; 1K8K [49]; 1IZN [61]; 1MIX [83]; 1MIZ [83]; 1QKR [93]; 1TR2 [92]; 1ST6 [94].

Article Snippet: , , FERM domain/Integrin β3 tail fragment (739–743) Complex , Gallus gallus , 200–400 , 1.9 , 0.204 , 1MIZ.

Techniques:

Journal: Nature Communications

Article Title: Enzymatic biosynthesis and immobilization of polyprotein verified at the single-molecule level

doi: 10.1038/s41467-019-10696-x

Figure Lengend Snippet: The OaAEP1-dependent polyprotein immobilization verified by single-molecule AFM. a The schematic shows that OaAEP1 facilitates a covalent anchoring of (poly)protein on a glass surface as well as a specifically probing by an XDoc-functionalized AFM tip. b Representative curves of (Ub) n in the specific attachment showed a high detachment force from the Coh-XDoc interaction and more unfolding peaks. c , d The statistical analysis of the (Ub) n indicated the number of curves with a specific number of ubiquitin peaks. Most curves showed the number of Ub unfolding peaks between three and six (>90%). e The scatter plot shows the relationship between unfolding/rupture force and ΔLc for (Ub) n , Xmodule of the XDoc (ΔLc = 35 nm), and the CBM. f The overlay of all curves with CBM unfolding first show the complete unfolding of the polyprotein (Ub) 6 using the OaAEP1-dependent polyprotein immobilization method ( n = 52). The covalent immobilization with a high detachment force enables the complete unfolding of all subdomains in the polyprotein. Source data of Fig. are provided as a Source Data file

Article Snippet: The gene coding for protein of interest: ubiquitin from human (Ub), rubredoxins (Rd, Rd represents Zn-formed rubredoxin from Clostridium pasteurianum , if not specified, Fe-form rubredoxin from Pyrococcus furiosus ), the B1 domain of immunoglobulin G (GB1), the cellulose-binding module (CBM), type III cohesin-dockerin-X module domain complex from Ruminococcus flavefaciens (Coh-Xmodule-Doc, or Coh-XDoc), tobacco etch virus (TEV) protease (fused with superfold GFP as GFP-TEV for use), elastin-like polypeptides (ELP) were ordered from Genscript Inc, respectively.

Techniques: Ubiquitin Proteomics