site-specific recombination Thermo Fisher Search Results


  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 95
    Millipore human thrombin
    Human Thrombin, supplied by Millipore, used in various techniques. Bioz Stars score: 95/100, based on 1180 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/human thrombin/product/Millipore
    Average 95 stars, based on 1180 article reviews
    Price from $9.99 to $1999.99
    human thrombin - by Bioz Stars, 2020-02
    95/100 stars
      Buy from Supplier

    90
    Thermo Fisher gateway lr clonase ii enzyme mix
    Gateway Lr Clonase Ii Enzyme Mix, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 90/100, based on 4329 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/gateway lr clonase ii enzyme mix/product/Thermo Fisher
    Average 90 stars, based on 4329 article reviews
    Price from $9.99 to $1999.99
    gateway lr clonase ii enzyme mix - by Bioz Stars, 2020-02
    90/100 stars
      Buy from Supplier

    90
    Thermo Fisher gateway site specific recombination technology
    Gateway Site Specific Recombination Technology, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 90/100, based on 38 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/gateway site specific recombination technology/product/Thermo Fisher
    Average 90 stars, based on 38 article reviews
    Price from $9.99 to $1999.99
    gateway site specific recombination technology - by Bioz Stars, 2020-02
    90/100 stars
      Buy from Supplier

    91
    Thermo Fisher gateway invitrogen attb recombination sites
    Gateway Invitrogen Attb Recombination Sites, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 91/100, based on 62 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/gateway invitrogen attb recombination sites/product/Thermo Fisher
    Average 91 stars, based on 62 article reviews
    Price from $9.99 to $1999.99
    gateway invitrogen attb recombination sites - by Bioz Stars, 2020-02
    91/100 stars
      Buy from Supplier

    78
    Thermo Fisher gateway invitrogen site specific recombination system
    Gateway Invitrogen Site Specific Recombination System, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 78/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/gateway invitrogen site specific recombination system/product/Thermo Fisher
    Average 78 stars, based on 8 article reviews
    Price from $9.99 to $1999.99
    gateway invitrogen site specific recombination system - by Bioz Stars, 2020-02
    78/100 stars
      Buy from Supplier

    78
    Thermo Fisher gateway site specific recombination ssr cloning system
    Gateway Site Specific Recombination Ssr Cloning System, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 78/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/gateway site specific recombination ssr cloning system/product/Thermo Fisher
    Average 78 stars, based on 6 article reviews
    Price from $9.99 to $1999.99
    gateway site specific recombination ssr cloning system - by Bioz Stars, 2020-02
    78/100 stars
      Buy from Supplier

    78
    Thermo Fisher widely used site specific recombination ssr cloning system
    Widely Used Site Specific Recombination Ssr Cloning System, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 78/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/widely used site specific recombination ssr cloning system/product/Thermo Fisher
    Average 78 stars, based on 7 article reviews
    Price from $9.99 to $1999.99
    widely used site specific recombination ssr cloning system - by Bioz Stars, 2020-02
    78/100 stars
      Buy from Supplier

    83
    Thermo Fisher gateway site specific recombinational cloning technology
    Gateway Site Specific Recombinational Cloning Technology, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 83/100, based on 13 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/gateway site specific recombinational cloning technology/product/Thermo Fisher
    Average 83 stars, based on 13 article reviews
    Price from $9.99 to $1999.99
    gateway site specific recombinational cloning technology - by Bioz Stars, 2020-02
    83/100 stars
      Buy from Supplier

    78
    Thermo Fisher flp recombinase mediated homologous recombination system
    Flp Recombinase Mediated Homologous Recombination System, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 78/100, based on 10 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/flp recombinase mediated homologous recombination system/product/Thermo Fisher
    Average 78 stars, based on 10 article reviews
    Price from $9.99 to $1999.99
    flp recombinase mediated homologous recombination system - by Bioz Stars, 2020-02
    78/100 stars
      Buy from Supplier

    78
    Thermo Fisher pentrtm 1a
    Pentrtm 1a, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 78/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pentrtm 1a/product/Thermo Fisher
    Average 78 stars, based on 6 article reviews
    Price from $9.99 to $1999.99
    pentrtm 1a - by Bioz Stars, 2020-02
    78/100 stars
      Buy from Supplier

    79
    Thermo Fisher pdest6 3 vector
    Pdest6 3 Vector, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 79/100, based on 16 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pdest6 3 vector/product/Thermo Fisher
    Average 79 stars, based on 16 article reviews
    Price from $9.99 to $1999.99
    pdest6 3 vector - by Bioz Stars, 2020-02
    79/100 stars
      Buy from Supplier

    96
    Thermo Fisher pog44 flp recombinase plasmid
    Pog44 Flp Recombinase Plasmid, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 96/100, based on 27 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pog44 flp recombinase plasmid/product/Thermo Fisher
    Average 96 stars, based on 27 article reviews
    Price from $9.99 to $1999.99
    pog44 flp recombinase plasmid - by Bioz Stars, 2020-02
    96/100 stars
      Buy from Supplier

    90
    Thermo Fisher multisite gateway technology
    Multisite Gateway Technology, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 90/100, based on 484 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/multisite gateway technology/product/Thermo Fisher
    Average 90 stars, based on 484 article reviews
    Price from $9.99 to $1999.99
    multisite gateway technology - by Bioz Stars, 2020-02
    90/100 stars
      Buy from Supplier

    90
    Thermo Fisher plenti6 v5 dest vector
    Plenti6 V5 Dest Vector, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 90/100, based on 226 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/plenti6 v5 dest vector/product/Thermo Fisher
    Average 90 stars, based on 226 article reviews
    Price from $9.99 to $1999.99
    plenti6 v5 dest vector - by Bioz Stars, 2020-02
    90/100 stars
      Buy from Supplier

    78
    Thermo Fisher fak phosphorylation site specific antibodies
    Association of the FERM domain with <t>FAK/CAKβ</t> chimeras and FAK mutants. (A) Schematic illustration of FAK/CAKβ chimeras utilized in this analysis. (B) Lysates of cells expressing the chimeras FFF (lanes 1 to 4), CFC (lanes 5 to 8), CCC (lanes 9 to 12), and FCF (lanes 13 to 16) were incubated with GST (lanes 2, 6, 10, and 14), GST-N-FAK (lanes 3, 7, 11, and 15), or GST-N-CAKβ (lanes 4, 8, 12, and 16) immobilized to glutathione-agarose beads. Chimeric proteins that bound to the GST fusion proteins were detected by Western blotting with <t>KT3.</t> Lysate was directly analyzed as a control (lanes 1, 5, 9, and 13). (C) Lysates of cells expressing the chimeras FFF (lanes 1 and 2), FFC (lanes 3 and 4) CFC (lanes 5 and 6), and CFF (lanes 7 and 8) were incubated with GST-N-FAK (lanes 2, 4, 6, and 8) immobilized to glutathione-agarose beads, and bound protein was detected by Western blotting with KT3. Lysate was directly analyzed as a control (lanes 1, 3, 5, and 7). (D) Lysates of untransfected CE cells (lane 4) or cells expressing wild-type FAK (lanes 5) or the FAK mutant dl51-377 (lane 6) were incubated with GST-N-FAK, and bound FAK was detected by Western blotting with the BC4 polyclonal antiserum. Lysates were directly analyzed as a control (lanes 1 to 3). (E) Lysates of CE cells expressing FAK (lanes 1 to 3), FAKY397F (lanes 4 and 5), or FAKK454M (lanes 6 and 7) were incubated with GST-N-FAK, and bound FAK was detected by Western blotting with KT3. Lysates were directly analyzed by Western blotting as a control (lanes 1, 4, and 6). As a negative control for FAK binding, GST was used.
    Fak Phosphorylation Site Specific Antibodies, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 78/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/fak phosphorylation site specific antibodies/product/Thermo Fisher
    Average 78 stars, based on 7 article reviews
    Price from $9.99 to $1999.99
    fak phosphorylation site specific antibodies - by Bioz Stars, 2020-02
    78/100 stars
      Buy from Supplier

    90
    Thermo Fisher pog44 flp recombinase expression vector
    Association of the FERM domain with <t>FAK/CAKβ</t> chimeras and FAK mutants. (A) Schematic illustration of FAK/CAKβ chimeras utilized in this analysis. (B) Lysates of cells expressing the chimeras FFF (lanes 1 to 4), CFC (lanes 5 to 8), CCC (lanes 9 to 12), and FCF (lanes 13 to 16) were incubated with GST (lanes 2, 6, 10, and 14), GST-N-FAK (lanes 3, 7, 11, and 15), or GST-N-CAKβ (lanes 4, 8, 12, and 16) immobilized to glutathione-agarose beads. Chimeric proteins that bound to the GST fusion proteins were detected by Western blotting with <t>KT3.</t> Lysate was directly analyzed as a control (lanes 1, 5, 9, and 13). (C) Lysates of cells expressing the chimeras FFF (lanes 1 and 2), FFC (lanes 3 and 4) CFC (lanes 5 and 6), and CFF (lanes 7 and 8) were incubated with GST-N-FAK (lanes 2, 4, 6, and 8) immobilized to glutathione-agarose beads, and bound protein was detected by Western blotting with KT3. Lysate was directly analyzed as a control (lanes 1, 3, 5, and 7). (D) Lysates of untransfected CE cells (lane 4) or cells expressing wild-type FAK (lanes 5) or the FAK mutant dl51-377 (lane 6) were incubated with GST-N-FAK, and bound FAK was detected by Western blotting with the BC4 polyclonal antiserum. Lysates were directly analyzed as a control (lanes 1 to 3). (E) Lysates of CE cells expressing FAK (lanes 1 to 3), FAKY397F (lanes 4 and 5), or FAKK454M (lanes 6 and 7) were incubated with GST-N-FAK, and bound FAK was detected by Western blotting with KT3. Lysates were directly analyzed by Western blotting as a control (lanes 1, 4, and 6). As a negative control for FAK binding, GST was used.
    Pog44 Flp Recombinase Expression Vector, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 90/100, based on 232 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pog44 flp recombinase expression vector/product/Thermo Fisher
    Average 90 stars, based on 232 article reviews
    Price from $9.99 to $1999.99
    pog44 flp recombinase expression vector - by Bioz Stars, 2020-02
    90/100 stars
      Buy from Supplier

    Image Search Results


    Association of the FERM domain with FAK/CAKβ chimeras and FAK mutants. (A) Schematic illustration of FAK/CAKβ chimeras utilized in this analysis. (B) Lysates of cells expressing the chimeras FFF (lanes 1 to 4), CFC (lanes 5 to 8), CCC (lanes 9 to 12), and FCF (lanes 13 to 16) were incubated with GST (lanes 2, 6, 10, and 14), GST-N-FAK (lanes 3, 7, 11, and 15), or GST-N-CAKβ (lanes 4, 8, 12, and 16) immobilized to glutathione-agarose beads. Chimeric proteins that bound to the GST fusion proteins were detected by Western blotting with KT3. Lysate was directly analyzed as a control (lanes 1, 5, 9, and 13). (C) Lysates of cells expressing the chimeras FFF (lanes 1 and 2), FFC (lanes 3 and 4) CFC (lanes 5 and 6), and CFF (lanes 7 and 8) were incubated with GST-N-FAK (lanes 2, 4, 6, and 8) immobilized to glutathione-agarose beads, and bound protein was detected by Western blotting with KT3. Lysate was directly analyzed as a control (lanes 1, 3, 5, and 7). (D) Lysates of untransfected CE cells (lane 4) or cells expressing wild-type FAK (lanes 5) or the FAK mutant dl51-377 (lane 6) were incubated with GST-N-FAK, and bound FAK was detected by Western blotting with the BC4 polyclonal antiserum. Lysates were directly analyzed as a control (lanes 1 to 3). (E) Lysates of CE cells expressing FAK (lanes 1 to 3), FAKY397F (lanes 4 and 5), or FAKK454M (lanes 6 and 7) were incubated with GST-N-FAK, and bound FAK was detected by Western blotting with KT3. Lysates were directly analyzed by Western blotting as a control (lanes 1, 4, and 6). As a negative control for FAK binding, GST was used.

    Journal: Molecular and Cellular Biology

    Article Title: FERM Domain Interaction Promotes FAK Signaling

    doi: 10.1128/MCB.24.12.5353-5368.2004

    Figure Lengend Snippet: Association of the FERM domain with FAK/CAKβ chimeras and FAK mutants. (A) Schematic illustration of FAK/CAKβ chimeras utilized in this analysis. (B) Lysates of cells expressing the chimeras FFF (lanes 1 to 4), CFC (lanes 5 to 8), CCC (lanes 9 to 12), and FCF (lanes 13 to 16) were incubated with GST (lanes 2, 6, 10, and 14), GST-N-FAK (lanes 3, 7, 11, and 15), or GST-N-CAKβ (lanes 4, 8, 12, and 16) immobilized to glutathione-agarose beads. Chimeric proteins that bound to the GST fusion proteins were detected by Western blotting with KT3. Lysate was directly analyzed as a control (lanes 1, 5, 9, and 13). (C) Lysates of cells expressing the chimeras FFF (lanes 1 and 2), FFC (lanes 3 and 4) CFC (lanes 5 and 6), and CFF (lanes 7 and 8) were incubated with GST-N-FAK (lanes 2, 4, 6, and 8) immobilized to glutathione-agarose beads, and bound protein was detected by Western blotting with KT3. Lysate was directly analyzed as a control (lanes 1, 3, 5, and 7). (D) Lysates of untransfected CE cells (lane 4) or cells expressing wild-type FAK (lanes 5) or the FAK mutant dl51-377 (lane 6) were incubated with GST-N-FAK, and bound FAK was detected by Western blotting with the BC4 polyclonal antiserum. Lysates were directly analyzed as a control (lanes 1 to 3). (E) Lysates of CE cells expressing FAK (lanes 1 to 3), FAKY397F (lanes 4 and 5), or FAKK454M (lanes 6 and 7) were incubated with GST-N-FAK, and bound FAK was detected by Western blotting with KT3. Lysates were directly analyzed by Western blotting as a control (lanes 1, 4, and 6). As a negative control for FAK binding, GST was used.

    Article Snippet: The KT3 monoclonal antibody (Covance, Princeton, N.J.), FAK phosphorylation site-specific antibodies (Biosource International, Camarillo, Calif.), a Pyk2/CAKβ monoclonal antibody and the 4G10 phosphotyrosine antibody (Upstate USA Inc., Lake Placid, N.Y.), and the RC20 phosphotyrosine antibody (BD Biosciences, San Diego, Calif.) were purchased commercially.

    Techniques: Expressing, Field Flow Fractionation, Countercurrent Chromatography, Incubation, Western Blot, Mutagenesis, Negative Control, Binding Assay

    The N-terminal domain of FAK associates with FAK in vitro. (A) Epitope-tagged FAK (lanes 1 to 4) or CAKβ (lanes 5 to 8) was expressed in CE cells. Cell lysates (1 mg) were precleared with GST bound to glutathione beads and then incubated with 25 μg of GST, GST-N-FAK, or GST-N-CAKβ. Bound proteins were detected by Western blotting with KT3. Cell lysate (25 μg) was analyzed directly by Western blotting as a control (lanes 1 and 5). (B) Rat-1 cells (lanes 1 to 4) or CE cells (lanes 5 to 8) were lysed, precleared with GST, and then incubated with GST, GST-N-FAK, or GST-N-CAKβ. Endogenous FAK bound to the beads was detected by Western blotting with BC4. Lysate (25 μg) was directly blotted as a control (lanes 1 and 5). (C) The blot in panel B was stripped and reprobed with a CAKβ monoclonal antibody to examine endogenous CAKβ binding to the GST fusion proteins. Note that CE cells express no endogenous CAKβ, and these samples were excluded from the analysis. (D) A fraction of the products from the GST pulldowns performed on Rat-1 cell lysates (B and C) were analyzed by SDS-PAGE andCoomassie blue staining to demonstrate that equal amounts of each fusion protein was used in the analysis. (E) The FAK N-terminal domain was transiently coexpressed with full-length FAK in 293 cells. Expression of each was determined by Western blotting of 25 μg of lysate with monoclonal antibody 4.47 (lanes 1 to 3). Full-length FAK was immunoprecipitated (IP) with monoclonal antibody 2A7, and the immunoprecipitated FAK and coimmunoprecipitating N-terminal domain were detected by Western blotting with monoclonal antibody 4.47 (lanes 7 to 9). As a specificity control for coimmunoprecipitation, mock precipitations with protein G alone were also performed (lanes 4 to 6). Numbers on the left are molecular weights in thousands.

    Journal: Molecular and Cellular Biology

    Article Title: FERM Domain Interaction Promotes FAK Signaling

    doi: 10.1128/MCB.24.12.5353-5368.2004

    Figure Lengend Snippet: The N-terminal domain of FAK associates with FAK in vitro. (A) Epitope-tagged FAK (lanes 1 to 4) or CAKβ (lanes 5 to 8) was expressed in CE cells. Cell lysates (1 mg) were precleared with GST bound to glutathione beads and then incubated with 25 μg of GST, GST-N-FAK, or GST-N-CAKβ. Bound proteins were detected by Western blotting with KT3. Cell lysate (25 μg) was analyzed directly by Western blotting as a control (lanes 1 and 5). (B) Rat-1 cells (lanes 1 to 4) or CE cells (lanes 5 to 8) were lysed, precleared with GST, and then incubated with GST, GST-N-FAK, or GST-N-CAKβ. Endogenous FAK bound to the beads was detected by Western blotting with BC4. Lysate (25 μg) was directly blotted as a control (lanes 1 and 5). (C) The blot in panel B was stripped and reprobed with a CAKβ monoclonal antibody to examine endogenous CAKβ binding to the GST fusion proteins. Note that CE cells express no endogenous CAKβ, and these samples were excluded from the analysis. (D) A fraction of the products from the GST pulldowns performed on Rat-1 cell lysates (B and C) were analyzed by SDS-PAGE andCoomassie blue staining to demonstrate that equal amounts of each fusion protein was used in the analysis. (E) The FAK N-terminal domain was transiently coexpressed with full-length FAK in 293 cells. Expression of each was determined by Western blotting of 25 μg of lysate with monoclonal antibody 4.47 (lanes 1 to 3). Full-length FAK was immunoprecipitated (IP) with monoclonal antibody 2A7, and the immunoprecipitated FAK and coimmunoprecipitating N-terminal domain were detected by Western blotting with monoclonal antibody 4.47 (lanes 7 to 9). As a specificity control for coimmunoprecipitation, mock precipitations with protein G alone were also performed (lanes 4 to 6). Numbers on the left are molecular weights in thousands.

    Article Snippet: The KT3 monoclonal antibody (Covance, Princeton, N.J.), FAK phosphorylation site-specific antibodies (Biosource International, Camarillo, Calif.), a Pyk2/CAKβ monoclonal antibody and the 4G10 phosphotyrosine antibody (Upstate USA Inc., Lake Placid, N.Y.), and the RC20 phosphotyrosine antibody (BD Biosciences, San Diego, Calif.) were purchased commercially.

    Techniques: In Vitro, Incubation, Western Blot, Binding Assay, SDS Page, Staining, Expressing, Immunoprecipitation

    Model of the FERM domain of FAK. (A) The backbone of the model of the FERM domain of FAK is shown. This model contains FAK residues 60 to 349. The ubiquitin-like subdomain (F1), acyl-CoA binding protein (BP)-like subdomain (F2) and PH/PTB/EVH-like subdomain (F3) are indicated. (B) The surface of the model of the FERM domain is shown. In the top panel, electrostatic potential is indicated colorimetrically, with red indicating negative potential and blue indicating positive potential. In the middle panel, the surface of the FERM domain is shown with residues that are identical from Drosophila to human (green) and highly conserved residues (black) indicated. In the bottom panel, the surface of the FERM domain is shown with residues that are identical between FAK and CAKβ in yellow. The circled region indicates a highly conserved basic patch at the tip of the F2 subdomain.

    Journal: Molecular and Cellular Biology

    Article Title: FERM Domain Interaction Promotes FAK Signaling

    doi: 10.1128/MCB.24.12.5353-5368.2004

    Figure Lengend Snippet: Model of the FERM domain of FAK. (A) The backbone of the model of the FERM domain of FAK is shown. This model contains FAK residues 60 to 349. The ubiquitin-like subdomain (F1), acyl-CoA binding protein (BP)-like subdomain (F2) and PH/PTB/EVH-like subdomain (F3) are indicated. (B) The surface of the model of the FERM domain is shown. In the top panel, electrostatic potential is indicated colorimetrically, with red indicating negative potential and blue indicating positive potential. In the middle panel, the surface of the FERM domain is shown with residues that are identical from Drosophila to human (green) and highly conserved residues (black) indicated. In the bottom panel, the surface of the FERM domain is shown with residues that are identical between FAK and CAKβ in yellow. The circled region indicates a highly conserved basic patch at the tip of the F2 subdomain.

    Article Snippet: The KT3 monoclonal antibody (Covance, Princeton, N.J.), FAK phosphorylation site-specific antibodies (Biosource International, Camarillo, Calif.), a Pyk2/CAKβ monoclonal antibody and the 4G10 phosphotyrosine antibody (Upstate USA Inc., Lake Placid, N.Y.), and the RC20 phosphotyrosine antibody (BD Biosciences, San Diego, Calif.) were purchased commercially.

    Techniques: Binding Assay

    Point mutations in the FERM domain disrupt the interaction with FAK. (A) Lysates of CE cells overexpressing epitope-tagged FAK were precleared with GST and then incubated with GST (lane 2), GST-N-FAK (lane 3), GST-KAKTLR (lane 4), or GST-KK (lane 5). Bound FAK was detected by Western blotting with KT3. Lysate (25 μg) was analyzed as a control (lane 1). (B) As a loading control, a fraction of product from each pulldown from panel A was analyzed by SDS-PAGE and Coomassie blue staining. (C) The wild-type (WT) FERM domain (lane 1) or FERM domain with the KAKTLR mutation (lane 2) was transiently coexpressed with full-length, wild-type FAK in HEK 293 cells. FAK was immunoprecipitated with 2A7, and the immune complexes were analyzed by Western blotting with 4.47 (top panel). Lysates were blotted with 4.47 to verify equal expression of the FERM domains (bottom panel). Numbers on the left are molecular weights in thousands. (D) Purified recombinant catalytic domain of FAK was incubated with GST-N-FAK (lanes 2 and 3), GST-KAKTLR (lanes 5 and 6), or GST (lanes 8 and 9) immobilized on glutathione beads in the presence of Mg 2+ and ATP. After washing and elution with free glutathione, samples were analyzed by Western blotting for phosphotyrosine (pTyr) (top panel) or by SDS-PAGE and Coomassie blue staining (bottom panel). As controls, GST fusion proteins were also incubated in the absence of the purified recombinant catalytic domain (lanes 1, 4, and 7).

    Journal: Molecular and Cellular Biology

    Article Title: FERM Domain Interaction Promotes FAK Signaling

    doi: 10.1128/MCB.24.12.5353-5368.2004

    Figure Lengend Snippet: Point mutations in the FERM domain disrupt the interaction with FAK. (A) Lysates of CE cells overexpressing epitope-tagged FAK were precleared with GST and then incubated with GST (lane 2), GST-N-FAK (lane 3), GST-KAKTLR (lane 4), or GST-KK (lane 5). Bound FAK was detected by Western blotting with KT3. Lysate (25 μg) was analyzed as a control (lane 1). (B) As a loading control, a fraction of product from each pulldown from panel A was analyzed by SDS-PAGE and Coomassie blue staining. (C) The wild-type (WT) FERM domain (lane 1) or FERM domain with the KAKTLR mutation (lane 2) was transiently coexpressed with full-length, wild-type FAK in HEK 293 cells. FAK was immunoprecipitated with 2A7, and the immune complexes were analyzed by Western blotting with 4.47 (top panel). Lysates were blotted with 4.47 to verify equal expression of the FERM domains (bottom panel). Numbers on the left are molecular weights in thousands. (D) Purified recombinant catalytic domain of FAK was incubated with GST-N-FAK (lanes 2 and 3), GST-KAKTLR (lanes 5 and 6), or GST (lanes 8 and 9) immobilized on glutathione beads in the presence of Mg 2+ and ATP. After washing and elution with free glutathione, samples were analyzed by Western blotting for phosphotyrosine (pTyr) (top panel) or by SDS-PAGE and Coomassie blue staining (bottom panel). As controls, GST fusion proteins were also incubated in the absence of the purified recombinant catalytic domain (lanes 1, 4, and 7).

    Article Snippet: The KT3 monoclonal antibody (Covance, Princeton, N.J.), FAK phosphorylation site-specific antibodies (Biosource International, Camarillo, Calif.), a Pyk2/CAKβ monoclonal antibody and the 4G10 phosphotyrosine antibody (Upstate USA Inc., Lake Placid, N.Y.), and the RC20 phosphotyrosine antibody (BD Biosciences, San Diego, Calif.) were purchased commercially.

    Techniques: Incubation, Western Blot, SDS Page, Staining, Mutagenesis, Immunoprecipitation, Expressing, Purification, Recombinant