mass spectrometry data processing program peaks Search Results


superose 6  (GE Healthcare)
94
GE Healthcare superose 6
Preparation of purified recombinant human coatomer . (A) Schematic diagram showing the domain organization of each subunits of human coatomer. The available atomic structures with their PDB IDs and sequence coverage are also labeled. (B) Coomassie-blue stained SDS-PAGE of recombinant coatomer with all the subunits labeled. Asterisk indicates the N-terminal fragment of δ-COP, which is confirmed by mass spectrometry and Western blot. (C) Gel filtration of purified coatomer. The retention volume of the single peak on the column of <t>Superose</t> <t>6</t> (GE healthcare) is 11.4 mL. (D) Native PAGE of the purified coatomer. Arrow indicates the intact coatomer
Superose 6, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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tandem mass spectrometry ms/ms  (Micromass UK Limited)
90
Micromass UK Limited tandem mass spectrometry ms/ms
Preparation of purified recombinant human coatomer . (A) Schematic diagram showing the domain organization of each subunits of human coatomer. The available atomic structures with their PDB IDs and sequence coverage are also labeled. (B) Coomassie-blue stained SDS-PAGE of recombinant coatomer with all the subunits labeled. Asterisk indicates the N-terminal fragment of δ-COP, which is confirmed by mass spectrometry and Western blot. (C) Gel filtration of purified coatomer. The retention volume of the single peak on the column of <t>Superose</t> <t>6</t> (GE healthcare) is 11.4 mL. (D) Native PAGE of the purified coatomer. Arrow indicates the intact coatomer
Tandem Mass Spectrometry Ms/Ms, supplied by Micromass UK Limited, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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tandem mass spectrometry ms/ms - by Bioz Stars, 2026-07
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maldi ms tofspec 2e  (Micromass UK Limited)
90
Micromass UK Limited maldi ms tofspec 2e
Preparation of purified recombinant human coatomer . (A) Schematic diagram showing the domain organization of each subunits of human coatomer. The available atomic structures with their PDB IDs and sequence coverage are also labeled. (B) Coomassie-blue stained SDS-PAGE of recombinant coatomer with all the subunits labeled. Asterisk indicates the N-terminal fragment of δ-COP, which is confirmed by mass spectrometry and Western blot. (C) Gel filtration of purified coatomer. The retention volume of the single peak on the column of <t>Superose</t> <t>6</t> (GE healthcare) is 11.4 mL. (D) Native PAGE of the purified coatomer. Arrow indicates the intact coatomer
Maldi Ms Tofspec 2e, supplied by Micromass UK Limited, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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nist mass spectrometry database  (Federation of European Neuroscience Societies)
90
Federation of European Neuroscience Societies nist mass spectrometry database
Preparation of purified recombinant human coatomer . (A) Schematic diagram showing the domain organization of each subunits of human coatomer. The available atomic structures with their PDB IDs and sequence coverage are also labeled. (B) Coomassie-blue stained SDS-PAGE of recombinant coatomer with all the subunits labeled. Asterisk indicates the N-terminal fragment of δ-COP, which is confirmed by mass spectrometry and Western blot. (C) Gel filtration of purified coatomer. The retention volume of the single peak on the column of <t>Superose</t> <t>6</t> (GE healthcare) is 11.4 mL. (D) Native PAGE of the purified coatomer. Arrow indicates the intact coatomer
Nist Mass Spectrometry Database, supplied by Federation of European Neuroscience Societies, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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package maldi biotypertm 3 0  (Bruker Corporation)
99
Bruker Corporation package maldi biotypertm 3 0
Preparation of purified recombinant human coatomer . (A) Schematic diagram showing the domain organization of each subunits of human coatomer. The available atomic structures with their PDB IDs and sequence coverage are also labeled. (B) Coomassie-blue stained SDS-PAGE of recombinant coatomer with all the subunits labeled. Asterisk indicates the N-terminal fragment of δ-COP, which is confirmed by mass spectrometry and Western blot. (C) Gel filtration of purified coatomer. The retention volume of the single peak on the column of <t>Superose</t> <t>6</t> (GE healthcare) is 11.4 mL. (D) Native PAGE of the purified coatomer. Arrow indicates the intact coatomer
Package Maldi Biotypertm 3 0, supplied by Bruker Corporation, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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package maldi biotypertm 3 0 - by Bioz Stars, 2026-07
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mass spectrometry libraries  (National Institute of Standards and Technology)
90
National Institute of Standards and Technology mass spectrometry libraries
Preparation of purified recombinant human coatomer . (A) Schematic diagram showing the domain organization of each subunits of human coatomer. The available atomic structures with their PDB IDs and sequence coverage are also labeled. (B) Coomassie-blue stained SDS-PAGE of recombinant coatomer with all the subunits labeled. Asterisk indicates the N-terminal fragment of δ-COP, which is confirmed by mass spectrometry and Western blot. (C) Gel filtration of purified coatomer. The retention volume of the single peak on the column of <t>Superose</t> <t>6</t> (GE healthcare) is 11.4 mL. (D) Native PAGE of the purified coatomer. Arrow indicates the intact coatomer
Mass Spectrometry Libraries, supplied by National Institute of Standards and Technology, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/mass+spectrometry+data+processing+program+peaks/pmc11381841-74-13-18?v=National+Institute+of+Standards+and+Technology
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mass spectrometry libraries - by Bioz Stars, 2026-07
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lc ms data  (Shimadzu Corporation)
99
Shimadzu Corporation lc ms data
Preparation of purified recombinant human coatomer . (A) Schematic diagram showing the domain organization of each subunits of human coatomer. The available atomic structures with their PDB IDs and sequence coverage are also labeled. (B) Coomassie-blue stained SDS-PAGE of recombinant coatomer with all the subunits labeled. Asterisk indicates the N-terminal fragment of δ-COP, which is confirmed by mass spectrometry and Western blot. (C) Gel filtration of purified coatomer. The retention volume of the single peak on the column of <t>Superose</t> <t>6</t> (GE healthcare) is 11.4 mL. (D) Native PAGE of the purified coatomer. Arrow indicates the intact coatomer
Lc Ms Data, supplied by Shimadzu Corporation, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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lc ms data - by Bioz Stars, 2026-07
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sgk269 antibodies  (Santa Cruz Biotechnology)
93
Santa Cruz Biotechnology sgk269 antibodies
FIGURE 3. Identification of novel <t>SgK269</t> binding partners. a, schematic of MS-based proteomics workflow. Anti-FLAG IPs prepared from MCF-10A cells expressing FLAG-tagged SgK269 were subjected to on-bead tryptic digestion. Resulting peptides were subjected to LC-MS/MS analysis. b, interactome of SgK269 represented as a volcano plot of the MS data. Proteins significantly enriched in the SgK269 IP with an adjusted p value 0.01 and a fold change of 2 against the control IP are displayed as red dots.
Sgk269 Antibodies, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/mass+spectrometry+data+processing+program+peaks/10__1074_slash_jbc__m116__748897-194-17-25?v=Santa+Cruz+Biotechnology
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sgk269 antibodies - by Bioz Stars, 2026-07
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peaks mass spectrometry software  (Bioinformatics Solutions Inc)
86
Bioinformatics Solutions Inc peaks mass spectrometry software
FIGURE 3. Identification of novel <t>SgK269</t> binding partners. a, schematic of MS-based proteomics workflow. Anti-FLAG IPs prepared from MCF-10A cells expressing FLAG-tagged SgK269 were subjected to on-bead tryptic digestion. Resulting peptides were subjected to LC-MS/MS analysis. b, interactome of SgK269 represented as a volcano plot of the MS data. Proteins significantly enriched in the SgK269 IP with an adjusted p value 0.01 and a fold change of 2 against the control IP are displayed as red dots.
Peaks Mass Spectrometry Software, supplied by Bioinformatics Solutions Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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peaks mass spectrometry software - by Bioz Stars, 2026-07
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de-str madli-tof mass spectrometer  (PerSeptive Biosystems Inc)
90
PerSeptive Biosystems Inc de-str madli-tof mass spectrometer
FIGURE 3. Identification of novel <t>SgK269</t> binding partners. a, schematic of MS-based proteomics workflow. Anti-FLAG IPs prepared from MCF-10A cells expressing FLAG-tagged SgK269 were subjected to on-bead tryptic digestion. Resulting peptides were subjected to LC-MS/MS analysis. b, interactome of SgK269 represented as a volcano plot of the MS data. Proteins significantly enriched in the SgK269 IP with an adjusted p value 0.01 and a fold change of 2 against the control IP are displayed as red dots.
De Str Madli Tof Mass Spectrometer, supplied by PerSeptive Biosystems Inc, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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quattro lcz tandem mass spectrometer  (Micromass UK Limited)
90
Micromass UK Limited quattro lcz tandem mass spectrometer
FIGURE 3. Identification of novel <t>SgK269</t> binding partners. a, schematic of MS-based proteomics workflow. Anti-FLAG IPs prepared from MCF-10A cells expressing FLAG-tagged SgK269 were subjected to on-bead tryptic digestion. Resulting peptides were subjected to LC-MS/MS analysis. b, interactome of SgK269 represented as a volcano plot of the MS data. Proteins significantly enriched in the SgK269 IP with an adjusted p value 0.01 and a fold change of 2 against the control IP are displayed as red dots.
Quattro Lcz Tandem Mass Spectrometer, supplied by Micromass UK Limited, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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isotope ratio mass spectrometer irms  (Finnigan Corporation)
86
Finnigan Corporation isotope ratio mass spectrometer irms
FIGURE 3. Identification of novel <t>SgK269</t> binding partners. a, schematic of MS-based proteomics workflow. Anti-FLAG IPs prepared from MCF-10A cells expressing FLAG-tagged SgK269 were subjected to on-bead tryptic digestion. Resulting peptides were subjected to LC-MS/MS analysis. b, interactome of SgK269 represented as a volcano plot of the MS data. Proteins significantly enriched in the SgK269 IP with an adjusted p value 0.01 and a fold change of 2 against the control IP are displayed as red dots.
Isotope Ratio Mass Spectrometer Irms, supplied by Finnigan Corporation, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Image Search Results


Preparation of purified recombinant human coatomer . (A) Schematic diagram showing the domain organization of each subunits of human coatomer. The available atomic structures with their PDB IDs and sequence coverage are also labeled. (B) Coomassie-blue stained SDS-PAGE of recombinant coatomer with all the subunits labeled. Asterisk indicates the N-terminal fragment of δ-COP, which is confirmed by mass spectrometry and Western blot. (C) Gel filtration of purified coatomer. The retention volume of the single peak on the column of Superose 6 (GE healthcare) is 11.4 mL. (D) Native PAGE of the purified coatomer. Arrow indicates the intact coatomer

Journal: Protein & Cell

Article Title: Structural characterization of coatomer in its cytosolic state

doi: 10.1007/s13238-016-0296-z

Figure Lengend Snippet: Preparation of purified recombinant human coatomer . (A) Schematic diagram showing the domain organization of each subunits of human coatomer. The available atomic structures with their PDB IDs and sequence coverage are also labeled. (B) Coomassie-blue stained SDS-PAGE of recombinant coatomer with all the subunits labeled. Asterisk indicates the N-terminal fragment of δ-COP, which is confirmed by mass spectrometry and Western blot. (C) Gel filtration of purified coatomer. The retention volume of the single peak on the column of Superose 6 (GE healthcare) is 11.4 mL. (D) Native PAGE of the purified coatomer. Arrow indicates the intact coatomer

Article Snippet: The retention volume of the single peak on the column of Superose 6 (GE healthcare) is 11.4 mL. (D) Native PAGE of the purified coatomer.

Techniques: Purification, Recombinant, Sequencing, Labeling, Staining, SDS Page, Mass Spectrometry, Western Blot, Filtration, Clear Native PAGE

FIGURE 3. Identification of novel SgK269 binding partners. a, schematic of MS-based proteomics workflow. Anti-FLAG IPs prepared from MCF-10A cells expressing FLAG-tagged SgK269 were subjected to on-bead tryptic digestion. Resulting peptides were subjected to LC-MS/MS analysis. b, interactome of SgK269 represented as a volcano plot of the MS data. Proteins significantly enriched in the SgK269 IP with an adjusted p value 0.01 and a fold change of 2 against the control IP are displayed as red dots.

Journal: Journal of Biological Chemistry

Article Title: Homo- and Heterotypic Association Regulates Signaling by the SgK269/PEAK1 and SgK223 Pseudokinases

doi: 10.1074/jbc.m116.748897

Figure Lengend Snippet: FIGURE 3. Identification of novel SgK269 binding partners. a, schematic of MS-based proteomics workflow. Anti-FLAG IPs prepared from MCF-10A cells expressing FLAG-tagged SgK269 were subjected to on-bead tryptic digestion. Resulting peptides were subjected to LC-MS/MS analysis. b, interactome of SgK269 represented as a volcano plot of the MS data. Proteins significantly enriched in the SgK269 IP with an adjusted p value 0.01 and a fold change of 2 against the control IP are displayed as red dots.

Article Snippet: -Actin (catalog no. sc-69879), normal rabbit IgG (catalog no. sc-2027), normal mouse IgG (catalog no. sc-2025), and SgK269 antibodies (catalog no. sc-100403) were purchased from Santa Cruz Biotechnology (Dallas, TX); HA (catalog. 11867423001) was purchased from Roche Applied Science (Mannheim, Ger- FIGURE 10.

Techniques: Binding Assay, Expressing, Liquid Chromatography with Mass Spectroscopy, Control

FIGURE 4. CH and PK domains are critical for heterotypic association of SgK269 and SgK223. a–c, SgK269 and SgK223 associate in vivo. Co-immunopre- cipitation of exogenous FLAG/HA-tagged SgK269 with endogenous SgK223 in MCF-10A cells (a) and endogenous SgK269 and SgK223 in MCF-10A cells (b) and MDA-MB-231 breast cancer cells (c) is shown. IgG was used as a negative control for b and c. d, schematic representation of SgK269 and SgK223 wild-type constructs containing N-terminal HA tags and C-terminal FLAG and Myc tags. Truncation mutants were generated for both, which lacked the N-terminal region (N), the C-terminal -helical region (CH), or the pseudokinase domain (PK). e and f, identification of domains required for heterotypic association of SgK269 and SgK223. MCF-10A cells stably overexpressing epitope-tagged versions of the indicated SgK269 proteins (e) or SgK223 proteins (f) were harvested, and lysates were subjected to immunoprecipitation with an anti-FLAG antibody. IPs were blotted for HA and SgK223 (e) or SgK269 (f). All data are representative of at least two independent experiments.

Journal: Journal of Biological Chemistry

Article Title: Homo- and Heterotypic Association Regulates Signaling by the SgK269/PEAK1 and SgK223 Pseudokinases

doi: 10.1074/jbc.m116.748897

Figure Lengend Snippet: FIGURE 4. CH and PK domains are critical for heterotypic association of SgK269 and SgK223. a–c, SgK269 and SgK223 associate in vivo. Co-immunopre- cipitation of exogenous FLAG/HA-tagged SgK269 with endogenous SgK223 in MCF-10A cells (a) and endogenous SgK269 and SgK223 in MCF-10A cells (b) and MDA-MB-231 breast cancer cells (c) is shown. IgG was used as a negative control for b and c. d, schematic representation of SgK269 and SgK223 wild-type constructs containing N-terminal HA tags and C-terminal FLAG and Myc tags. Truncation mutants were generated for both, which lacked the N-terminal region (N), the C-terminal -helical region (CH), or the pseudokinase domain (PK). e and f, identification of domains required for heterotypic association of SgK269 and SgK223. MCF-10A cells stably overexpressing epitope-tagged versions of the indicated SgK269 proteins (e) or SgK223 proteins (f) were harvested, and lysates were subjected to immunoprecipitation with an anti-FLAG antibody. IPs were blotted for HA and SgK223 (e) or SgK269 (f). All data are representative of at least two independent experiments.

Article Snippet: -Actin (catalog no. sc-69879), normal rabbit IgG (catalog no. sc-2027), normal mouse IgG (catalog no. sc-2025), and SgK269 antibodies (catalog no. sc-100403) were purchased from Santa Cruz Biotechnology (Dallas, TX); HA (catalog. 11867423001) was purchased from Roche Applied Science (Mannheim, Ger- FIGURE 10.

Techniques: In Vivo, Negative Control, Construct, Generated, Stable Transfection, Immunoprecipitation

FIGURE 5. CH and PK domains are critical for homotypic association of SgK269 and SgK223. a, schematic representation of wild-type SgK269 with either HA tag or FLAG/Myc tags. Deletion mutants were generated with FLAG/Myc tags only. The same was undertaken for SgK223. b and c, identification of domains required for homotypic association of SgK269 and SgK223. HEK293 cells were co-transfected with plasmids encoding HA-tagged SgK269 (HA-WT) in combi- nation with plasmids encoding FLAG/Myc-tagged SgK269 or different deletion mutants (b). Lysates were subject to immunoprecipitation with an anti-FLAG antibody and Western blotted anti-HA or anti-FLAG as indicated. c, same procedure was followed, but this time co-expressing HA-tagged SgK223 with FLAG/Myc-tagged SgK223 proteins. All data are representative of at least two independent experiments.

Journal: Journal of Biological Chemistry

Article Title: Homo- and Heterotypic Association Regulates Signaling by the SgK269/PEAK1 and SgK223 Pseudokinases

doi: 10.1074/jbc.m116.748897

Figure Lengend Snippet: FIGURE 5. CH and PK domains are critical for homotypic association of SgK269 and SgK223. a, schematic representation of wild-type SgK269 with either HA tag or FLAG/Myc tags. Deletion mutants were generated with FLAG/Myc tags only. The same was undertaken for SgK223. b and c, identification of domains required for homotypic association of SgK269 and SgK223. HEK293 cells were co-transfected with plasmids encoding HA-tagged SgK269 (HA-WT) in combi- nation with plasmids encoding FLAG/Myc-tagged SgK269 or different deletion mutants (b). Lysates were subject to immunoprecipitation with an anti-FLAG antibody and Western blotted anti-HA or anti-FLAG as indicated. c, same procedure was followed, but this time co-expressing HA-tagged SgK223 with FLAG/Myc-tagged SgK223 proteins. All data are representative of at least two independent experiments.

Article Snippet: -Actin (catalog no. sc-69879), normal rabbit IgG (catalog no. sc-2027), normal mouse IgG (catalog no. sc-2025), and SgK269 antibodies (catalog no. sc-100403) were purchased from Santa Cruz Biotechnology (Dallas, TX); HA (catalog. 11867423001) was purchased from Roche Applied Science (Mannheim, Ger- FIGURE 10.

Techniques: Generated, Transfection, Immunoprecipitation, Western Blot, Expressing

FIGURE 6. CH region is critical for homotypic and heterotypic association of recombinant SgK269 and SgK223 C-terminal regions. Equal amounts of SgK269 (a and b) or SgK223 (c) GST fusion proteins on beads were incu- bated with recombinant His-tagged SgK269 (b) or SgK223 (a and c) proteins, as indicated. Following washing, samples were analyzed either by Coomassie Blue staining (bottom panels) or by Western blotting with an anti-His tag anti- body (top panels). GST was used as a negative control. Numbers on left-hand side of panels indicate position of corresponding size markers (sizes in kDa). All data are representative of at least two independent experiments.

Journal: Journal of Biological Chemistry

Article Title: Homo- and Heterotypic Association Regulates Signaling by the SgK269/PEAK1 and SgK223 Pseudokinases

doi: 10.1074/jbc.m116.748897

Figure Lengend Snippet: FIGURE 6. CH region is critical for homotypic and heterotypic association of recombinant SgK269 and SgK223 C-terminal regions. Equal amounts of SgK269 (a and b) or SgK223 (c) GST fusion proteins on beads were incu- bated with recombinant His-tagged SgK269 (b) or SgK223 (a and c) proteins, as indicated. Following washing, samples were analyzed either by Coomassie Blue staining (bottom panels) or by Western blotting with an anti-His tag anti- body (top panels). GST was used as a negative control. Numbers on left-hand side of panels indicate position of corresponding size markers (sizes in kDa). All data are representative of at least two independent experiments.

Article Snippet: -Actin (catalog no. sc-69879), normal rabbit IgG (catalog no. sc-2027), normal mouse IgG (catalog no. sc-2025), and SgK269 antibodies (catalog no. sc-100403) were purchased from Santa Cruz Biotechnology (Dallas, TX); HA (catalog. 11867423001) was purchased from Roche Applied Science (Mannheim, Ger- FIGURE 10.

Techniques: Recombinant, Staining, Western Blot, Negative Control

FIGURE 7. Characterization of homotypic and heterotypic association of recombinant SgK269 and SgK223 C-terminal regions via size-exclusion chromatography. a, Coomassie staining of purified recombinant His-tagged SgK269 and SgK223 CH-PK and PK proteins used for size-exclusion chromatography. b and c, size-exclusion chromatography. Elution profile of SgK223-CH-PK was compared with SgK223-PK (b). Elution profile of SgK269-CH-PK was compared with SgK269-PK (c). Elution profiles of SgK269-CH-PK SgK223-CH-PK (d) and SgK269-CH-PK SgK223-PK (e) were compared with their individual elution profiles. The arrow highlights the shift in elution profile upon complex formation. Also shown are corresponding Coomassie-stained gel fractions highlighting complex formation (d) and its absence (e). All data are representative of at least three independent experiments.

Journal: Journal of Biological Chemistry

Article Title: Homo- and Heterotypic Association Regulates Signaling by the SgK269/PEAK1 and SgK223 Pseudokinases

doi: 10.1074/jbc.m116.748897

Figure Lengend Snippet: FIGURE 7. Characterization of homotypic and heterotypic association of recombinant SgK269 and SgK223 C-terminal regions via size-exclusion chromatography. a, Coomassie staining of purified recombinant His-tagged SgK269 and SgK223 CH-PK and PK proteins used for size-exclusion chromatography. b and c, size-exclusion chromatography. Elution profile of SgK223-CH-PK was compared with SgK223-PK (b). Elution profile of SgK269-CH-PK was compared with SgK269-PK (c). Elution profiles of SgK269-CH-PK SgK223-CH-PK (d) and SgK269-CH-PK SgK223-PK (e) were compared with their individual elution profiles. The arrow highlights the shift in elution profile upon complex formation. Also shown are corresponding Coomassie-stained gel fractions highlighting complex formation (d) and its absence (e). All data are representative of at least three independent experiments.

Article Snippet: -Actin (catalog no. sc-69879), normal rabbit IgG (catalog no. sc-2027), normal mouse IgG (catalog no. sc-2025), and SgK269 antibodies (catalog no. sc-100403) were purchased from Santa Cruz Biotechnology (Dallas, TX); HA (catalog. 11867423001) was purchased from Roche Applied Science (Mannheim, Ger- FIGURE 10.

Techniques: Recombinant, Size-exclusion Chromatography, Staining, Purification

FIGURE 8. Association of SgK223 with Grb2 is dependent on SgK269. MDA-MB-231 cells were subject to siRNA-mediated knockdown of SgK269. SgK223 and Grb2 IPs and cell lysates from control and SgK269 knockdown cells were subject to Western blotting as indicated. Data are representative of two independent experiments.

Journal: Journal of Biological Chemistry

Article Title: Homo- and Heterotypic Association Regulates Signaling by the SgK269/PEAK1 and SgK223 Pseudokinases

doi: 10.1074/jbc.m116.748897

Figure Lengend Snippet: FIGURE 8. Association of SgK223 with Grb2 is dependent on SgK269. MDA-MB-231 cells were subject to siRNA-mediated knockdown of SgK269. SgK223 and Grb2 IPs and cell lysates from control and SgK269 knockdown cells were subject to Western blotting as indicated. Data are representative of two independent experiments.

Article Snippet: -Actin (catalog no. sc-69879), normal rabbit IgG (catalog no. sc-2027), normal mouse IgG (catalog no. sc-2025), and SgK269 antibodies (catalog no. sc-100403) were purchased from Santa Cruz Biotechnology (Dallas, TX); HA (catalog. 11867423001) was purchased from Roche Applied Science (Mannheim, Ger- FIGURE 10.

Techniques: Knockdown, Control, Western Blot

FIGURE 9. Regulation of pseudokinase signal output by homo- and heterotypic association. a, characterization of SgK223 knock-out (KO) MCF-10A cells generated by CRISPR/Cas9. Western blotting was undertaken as indicated. b, relative migration rates of control (Ctrl) MCF-10A and SgK223 KO cells. Relative wound closure was measured using ImageJ analysis software, and results from three independent experiments are presented as a histogram. Data are expressed relative to the value for MCF-10A cells, which was arbitrarily set at 1.0. c, expression of SgK269 WT and deletion mutants in MCF-10A and SgK223 KO cells.Westernblottingwasundertakenasindicated.d,effectofSgK269oncellmigrationinMCF-10AandSgK223KOcells.Relativewoundclosurewasassessed as in b and is expressed relative to cells transfected with empty vector (DsR), which was arbitrarily set at 1.0. Data represent mean S.E., **, p 0.01; #, p 0.05. e, effect of SgK269 WT and deletion mutants on cell migration in SgK223 KO cells. Data represent mean S.E., **, p 0.01; ***, p 0.001; #, p 0.05.

Journal: Journal of Biological Chemistry

Article Title: Homo- and Heterotypic Association Regulates Signaling by the SgK269/PEAK1 and SgK223 Pseudokinases

doi: 10.1074/jbc.m116.748897

Figure Lengend Snippet: FIGURE 9. Regulation of pseudokinase signal output by homo- and heterotypic association. a, characterization of SgK223 knock-out (KO) MCF-10A cells generated by CRISPR/Cas9. Western blotting was undertaken as indicated. b, relative migration rates of control (Ctrl) MCF-10A and SgK223 KO cells. Relative wound closure was measured using ImageJ analysis software, and results from three independent experiments are presented as a histogram. Data are expressed relative to the value for MCF-10A cells, which was arbitrarily set at 1.0. c, expression of SgK269 WT and deletion mutants in MCF-10A and SgK223 KO cells.Westernblottingwasundertakenasindicated.d,effectofSgK269oncellmigrationinMCF-10AandSgK223KOcells.Relativewoundclosurewasassessed as in b and is expressed relative to cells transfected with empty vector (DsR), which was arbitrarily set at 1.0. Data represent mean S.E., **, p 0.01; #, p 0.05. e, effect of SgK269 WT and deletion mutants on cell migration in SgK223 KO cells. Data represent mean S.E., **, p 0.01; ***, p 0.001; #, p 0.05.

Article Snippet: -Actin (catalog no. sc-69879), normal rabbit IgG (catalog no. sc-2027), normal mouse IgG (catalog no. sc-2025), and SgK269 antibodies (catalog no. sc-100403) were purchased from Santa Cruz Biotechnology (Dallas, TX); HA (catalog. 11867423001) was purchased from Roche Applied Science (Mannheim, Ger- FIGURE 10.

Techniques: Knock-Out, Generated, CRISPR, Western Blot, Migration, Control, Software, Expressing, Transfection, Plasmid Preparation

FIGURE 10. Enhancement of Stat3 activation by SgK269 requires SgK223. a and b, SgK269 and SgK223 promote Stat3 activation in MCF-10A cells in a CH and PK domain-dependent manner. Lysates from MCF-10A cells expressing SgK269 (a) or SgK223 (b) WT or deletion mutants were subject to Western blotting analysis as indicated. c, SgK269 fails to enhance Stat3 activation in SgK223 KO MCF-10A cells. The panels show Western blotting of lysates prepared from two individual SgK223 KO cell lines expressing SgK269 WT or deletion mutants. d, knockdown of SgK223 in SgK269-overexpressing MCF-10A cells reduces Stat3 tyrosine phosphorylation. Lysates from cells transfected with control siRNA or two independent SgK223-selective siRNAs were Western blotted as shown. All data representative of at least two independent experiments.

Journal: Journal of Biological Chemistry

Article Title: Homo- and Heterotypic Association Regulates Signaling by the SgK269/PEAK1 and SgK223 Pseudokinases

doi: 10.1074/jbc.m116.748897

Figure Lengend Snippet: FIGURE 10. Enhancement of Stat3 activation by SgK269 requires SgK223. a and b, SgK269 and SgK223 promote Stat3 activation in MCF-10A cells in a CH and PK domain-dependent manner. Lysates from MCF-10A cells expressing SgK269 (a) or SgK223 (b) WT or deletion mutants were subject to Western blotting analysis as indicated. c, SgK269 fails to enhance Stat3 activation in SgK223 KO MCF-10A cells. The panels show Western blotting of lysates prepared from two individual SgK223 KO cell lines expressing SgK269 WT or deletion mutants. d, knockdown of SgK223 in SgK269-overexpressing MCF-10A cells reduces Stat3 tyrosine phosphorylation. Lysates from cells transfected with control siRNA or two independent SgK223-selective siRNAs were Western blotted as shown. All data representative of at least two independent experiments.

Article Snippet: -Actin (catalog no. sc-69879), normal rabbit IgG (catalog no. sc-2027), normal mouse IgG (catalog no. sc-2025), and SgK269 antibodies (catalog no. sc-100403) were purchased from Santa Cruz Biotechnology (Dallas, TX); HA (catalog. 11867423001) was purchased from Roche Applied Science (Mannheim, Ger- FIGURE 10.

Techniques: Activation Assay, Expressing, Western Blot, Knockdown, Phospho-proteomics, Transfection, Control

FIGURE 11. Homo- and heterotypic association of the SgK269 and SgK223 pseudokinase scaffolds as a mechanism to regulate signal output. a, proposed model. SgK269 and SgK223 recruit distinct repertoires of SH2 or PTB domain-containing signaling proteins to their contrasting complement of tyrosine phosphorylation sites. Reflecting this, homotypic and heterotypic complexes generate contrasting integrated signals. In this model, we have also proposed that the PK domains provide a selective binding interface upon interaction that differs between homotypic and heterotypic complexes. b, differen- tial expression of SgK269 and SgK223 among breast epithelial and breast cancer cell lines, as detected by Western blotting.

Journal: Journal of Biological Chemistry

Article Title: Homo- and Heterotypic Association Regulates Signaling by the SgK269/PEAK1 and SgK223 Pseudokinases

doi: 10.1074/jbc.m116.748897

Figure Lengend Snippet: FIGURE 11. Homo- and heterotypic association of the SgK269 and SgK223 pseudokinase scaffolds as a mechanism to regulate signal output. a, proposed model. SgK269 and SgK223 recruit distinct repertoires of SH2 or PTB domain-containing signaling proteins to their contrasting complement of tyrosine phosphorylation sites. Reflecting this, homotypic and heterotypic complexes generate contrasting integrated signals. In this model, we have also proposed that the PK domains provide a selective binding interface upon interaction that differs between homotypic and heterotypic complexes. b, differen- tial expression of SgK269 and SgK223 among breast epithelial and breast cancer cell lines, as detected by Western blotting.

Article Snippet: -Actin (catalog no. sc-69879), normal rabbit IgG (catalog no. sc-2027), normal mouse IgG (catalog no. sc-2025), and SgK269 antibodies (catalog no. sc-100403) were purchased from Santa Cruz Biotechnology (Dallas, TX); HA (catalog. 11867423001) was purchased from Roche Applied Science (Mannheim, Ger- FIGURE 10.

Techniques: Phospho-proteomics, Binding Assay, Expressing, Western Blot