The Experts below are selected from a list of 267 Experts worldwide ranked by ideXlab platform
Ron Bose - One of the best experts on this subject based on the ideXlab platform.
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Carboxyl Group footprinting mass spectrometry and molecular dynamics identify key interactions in the her2 her3 receptor tyrosine kinase interface
Journal of Biological Chemistry, 2013Co-Authors: Timothy S Collier, Wei Shen, John Monsey, Karthikeyan Diraviyam, David Sept, Ron BoseAbstract:The HER2 receptor tyrosine kinase is a driver oncogene in many human cancers, including breast and gastric cancer. Under physiologic levels of expression, HER2 heterodimerizes with other members of the EGF receptor/HER/ErbB family, and the HER2-HER3 dimer forms one of the most potent oncogenic receptor pairs. Previous structural biology studies have individually crystallized the kinase domains of HER2 and HER3, but the HER2-HER3 kinase domain heterodimer structure has yet to be solved. Using a reconstituted membrane system to form HER2-HER3 kinase domain heterodimers and Carboxyl Group footprinting mass spectrometry, we observed that HER2 and HER3 kinase domains preferentially form asymmetric heterodimers with HER3 and HER2 monomers occupying the donor and acceptor kinase positions, respectively. Conformational changes in the HER2 activation loop, as measured by changes in Carboxyl Group labeling, required both dimerization and nucleotide binding but did not require activation loop phosphorylation at Tyr-877. Molecular dynamics simulations on HER2-HER3 kinase dimers identify specific inter- and intramolecular interactions and were in good agreement with MS measurements. Specifically, several intermolecular ionic interactions between HER2 Lys-716-HER3 Glu-909, HER2 Glu-717-HER3 Lys-907, and HER2 Asp-871-HER3 Arg-948 were identified by molecular dynamics. We also evaluated the effect of the cancer-associated mutations HER2 D769H/D769Y, HER3 E909G, and HER3 R948K (also numbered HER3 E928G and R967K) on kinase activity in the context of this new structural model. This study provides valuable insights into the EGF receptor/HER/ErbB kinase structure and interactions, which can guide the design of future therapies.
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Carboxyl Group footprinting maps the dimerization interface and phosphorylation induced conformational changes of a membrane associated tyrosine kinase
Molecular & Cellular Proteomics, 2011Co-Authors: Hao Zhang, Michael L Gross, Wei Shen, Don L Rempel, John Monsey, Ilan Vidavsky, Ron BoseAbstract:: Her4 is a transmembrane receptor tyrosine kinase belonging to the ErbB-EGFR family. It plays a vital role in the cardiovascular and nervous systems, and mutations in Her4 have been found in melanoma and lung cancer. The kinase domain of Her4 forms a dimer complex, called the asymmetric dimer, which results in kinase activation. Although a crystal structure of the Her4 asymmetric dimer is known, the dimer affinity and the effect of the subsequent phosphorylation steps on kinase domain conformation are unknown. We report here the use of Carboxyl-Group footprinting MS on a recombinant expressed, Her4 kinase-domain construct to address these questions. Carboxyl-Group footprinting uses a water-soluble carbodiimide, 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide, in the presence of glycine ethyl ester, to modify accessible Carboxyl Groups on glutamate and aspartate residues. Comparisons of Her4 kinase-domain monomers versus dimers and of unphosphorylated versus phosphorylated dimers were made to map the dimerization interface and to determine phosphorylation induced-conformational changes. We detected 37 glutamate and aspartate residues that were modified, and we quantified their extents of modification by liquid chromatography MS. Five residues showed changes in Carboxyl-Group modification. Three of these residues are at the predicted dimer interface, as shown by the crystal structure, and the remaining two residues are on loops that likely have altered conformation in the kinase dimer. Incubating the Her4 kinase dimers with ATP resulted in dramatic increase in Tyr-850 phosphorylation, located on the activation loop, and this resulted in a conformational change in this loop, as evidenced by reduction in Carboxyl-Group modification. The kinase monomer-dimer equilibrium was measured using a titration format in which the extent of Carboxyl-Group footprinting was mathematically modeled to give the dimer association constant (1.5-6.8 × 10(12) dm(2)/mol). This suggests that the kinase-domain makes a significant contribution to the overall dimerization affinity of the full-length Her4 protein.
Hao Zhang - One of the best experts on this subject based on the ideXlab platform.
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isotope encoded Carboxyl Group footprinting for mass spectrometry based protein conformational studies
Journal of the American Society for Mass Spectrometry, 2016Co-Authors: Hao Zhang, Robert E Blankenship, Michael L GrossAbstract:We report an isotope-encoding method coupled with Carboxyl-Group footprinting to monitor protein conformational changes. The Carboxyl Groups of aspartic/glutamic acids and of the C-terminus of proteins can serve as reporters for protein conformational changes when labeled with glycine ethyl ester (GEE) mediated by carbodiimide. In the new development, isotope-encoded “heavy” and “light” GEE are used to label separately the two states of the orange carotenoid protein (OCP) from cyanobacteria. Two samples are mixed (1:1 ratio) and analyzed by a single LC-MS/MS experiment. The differences in labeling extent between the two states are represented by the ratio of the “heavy” and “light” peptides, providing information about protein conformational changes. Combining isotope-encoded MS quantitative analysis and Carboxyl-Group footprinting reduces the time of MS analysis and improves the sensitivity of GEE and other footprinting.
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Carboxyl Group footprinting maps the dimerization interface and phosphorylation induced conformational changes of a membrane associated tyrosine kinase
Molecular & Cellular Proteomics, 2011Co-Authors: Hao Zhang, Michael L Gross, Wei Shen, Don L Rempel, John Monsey, Ilan Vidavsky, Ron BoseAbstract:: Her4 is a transmembrane receptor tyrosine kinase belonging to the ErbB-EGFR family. It plays a vital role in the cardiovascular and nervous systems, and mutations in Her4 have been found in melanoma and lung cancer. The kinase domain of Her4 forms a dimer complex, called the asymmetric dimer, which results in kinase activation. Although a crystal structure of the Her4 asymmetric dimer is known, the dimer affinity and the effect of the subsequent phosphorylation steps on kinase domain conformation are unknown. We report here the use of Carboxyl-Group footprinting MS on a recombinant expressed, Her4 kinase-domain construct to address these questions. Carboxyl-Group footprinting uses a water-soluble carbodiimide, 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide, in the presence of glycine ethyl ester, to modify accessible Carboxyl Groups on glutamate and aspartate residues. Comparisons of Her4 kinase-domain monomers versus dimers and of unphosphorylated versus phosphorylated dimers were made to map the dimerization interface and to determine phosphorylation induced-conformational changes. We detected 37 glutamate and aspartate residues that were modified, and we quantified their extents of modification by liquid chromatography MS. Five residues showed changes in Carboxyl-Group modification. Three of these residues are at the predicted dimer interface, as shown by the crystal structure, and the remaining two residues are on loops that likely have altered conformation in the kinase dimer. Incubating the Her4 kinase dimers with ATP resulted in dramatic increase in Tyr-850 phosphorylation, located on the activation loop, and this resulted in a conformational change in this loop, as evidenced by reduction in Carboxyl-Group modification. The kinase monomer-dimer equilibrium was measured using a titration format in which the extent of Carboxyl-Group footprinting was mathematically modeled to give the dimer association constant (1.5-6.8 × 10(12) dm(2)/mol). This suggests that the kinase-domain makes a significant contribution to the overall dimerization affinity of the full-length Her4 protein.
Michael L Gross - One of the best experts on this subject based on the ideXlab platform.
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isotope encoded Carboxyl Group footprinting for mass spectrometry based protein conformational studies
Journal of the American Society for Mass Spectrometry, 2016Co-Authors: Hao Zhang, Robert E Blankenship, Michael L GrossAbstract:We report an isotope-encoding method coupled with Carboxyl-Group footprinting to monitor protein conformational changes. The Carboxyl Groups of aspartic/glutamic acids and of the C-terminus of proteins can serve as reporters for protein conformational changes when labeled with glycine ethyl ester (GEE) mediated by carbodiimide. In the new development, isotope-encoded “heavy” and “light” GEE are used to label separately the two states of the orange carotenoid protein (OCP) from cyanobacteria. Two samples are mixed (1:1 ratio) and analyzed by a single LC-MS/MS experiment. The differences in labeling extent between the two states are represented by the ratio of the “heavy” and “light” peptides, providing information about protein conformational changes. Combining isotope-encoded MS quantitative analysis and Carboxyl-Group footprinting reduces the time of MS analysis and improves the sensitivity of GEE and other footprinting.
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Carboxyl Group footprinting maps the dimerization interface and phosphorylation induced conformational changes of a membrane associated tyrosine kinase
Molecular & Cellular Proteomics, 2011Co-Authors: Hao Zhang, Michael L Gross, Wei Shen, Don L Rempel, John Monsey, Ilan Vidavsky, Ron BoseAbstract:: Her4 is a transmembrane receptor tyrosine kinase belonging to the ErbB-EGFR family. It plays a vital role in the cardiovascular and nervous systems, and mutations in Her4 have been found in melanoma and lung cancer. The kinase domain of Her4 forms a dimer complex, called the asymmetric dimer, which results in kinase activation. Although a crystal structure of the Her4 asymmetric dimer is known, the dimer affinity and the effect of the subsequent phosphorylation steps on kinase domain conformation are unknown. We report here the use of Carboxyl-Group footprinting MS on a recombinant expressed, Her4 kinase-domain construct to address these questions. Carboxyl-Group footprinting uses a water-soluble carbodiimide, 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide, in the presence of glycine ethyl ester, to modify accessible Carboxyl Groups on glutamate and aspartate residues. Comparisons of Her4 kinase-domain monomers versus dimers and of unphosphorylated versus phosphorylated dimers were made to map the dimerization interface and to determine phosphorylation induced-conformational changes. We detected 37 glutamate and aspartate residues that were modified, and we quantified their extents of modification by liquid chromatography MS. Five residues showed changes in Carboxyl-Group modification. Three of these residues are at the predicted dimer interface, as shown by the crystal structure, and the remaining two residues are on loops that likely have altered conformation in the kinase dimer. Incubating the Her4 kinase dimers with ATP resulted in dramatic increase in Tyr-850 phosphorylation, located on the activation loop, and this resulted in a conformational change in this loop, as evidenced by reduction in Carboxyl-Group modification. The kinase monomer-dimer equilibrium was measured using a titration format in which the extent of Carboxyl-Group footprinting was mathematically modeled to give the dimer association constant (1.5-6.8 × 10(12) dm(2)/mol). This suggests that the kinase-domain makes a significant contribution to the overall dimerization affinity of the full-length Her4 protein.
G M Chow - One of the best experts on this subject based on the ideXlab platform.
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Carboxyl Group co2h functionalized ferrimagnetic iron oxide nanoparticles for potential bio applications
Journal of Materials Chemistry, 2004Co-Authors: Shi Yu, G M ChowAbstract:A new approach to prepare surface-functionalized magnetic nanoparticles by synthesis of poly(methacrylic acid) (PMAA) coated maghemite nanoparticles in aqueous solution is reported. Maghemite (γ-Fe2O3) nanoparticles with an average diameter of 8 ± 2 nm were fabricated and subsequently coated with PMAA by emulsion polymerization. The FTIR study and thermal analysis confirmed the chemical adsorption of methacrylic acid on the maghemite nanoparticle surface, and suggested a symmetrical Carboxylate bonding. The free Carboxyl Group of PMAA, which was verified by FTIR spectroscopy and zeta potential measurement, provided the site for immobilization of foreign molecules. The PMAA coated maghemite nanoparticles were demonstrated as potential magnetically targeted drug carriers by adsorbing an anti-cancer drug (carboplatin) via the ion–dipole interaction between CO2− of PMAA and carboplatin.
John Monsey - One of the best experts on this subject based on the ideXlab platform.
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Carboxyl Group footprinting mass spectrometry and molecular dynamics identify key interactions in the her2 her3 receptor tyrosine kinase interface
Journal of Biological Chemistry, 2013Co-Authors: Timothy S Collier, Wei Shen, John Monsey, Karthikeyan Diraviyam, David Sept, Ron BoseAbstract:The HER2 receptor tyrosine kinase is a driver oncogene in many human cancers, including breast and gastric cancer. Under physiologic levels of expression, HER2 heterodimerizes with other members of the EGF receptor/HER/ErbB family, and the HER2-HER3 dimer forms one of the most potent oncogenic receptor pairs. Previous structural biology studies have individually crystallized the kinase domains of HER2 and HER3, but the HER2-HER3 kinase domain heterodimer structure has yet to be solved. Using a reconstituted membrane system to form HER2-HER3 kinase domain heterodimers and Carboxyl Group footprinting mass spectrometry, we observed that HER2 and HER3 kinase domains preferentially form asymmetric heterodimers with HER3 and HER2 monomers occupying the donor and acceptor kinase positions, respectively. Conformational changes in the HER2 activation loop, as measured by changes in Carboxyl Group labeling, required both dimerization and nucleotide binding but did not require activation loop phosphorylation at Tyr-877. Molecular dynamics simulations on HER2-HER3 kinase dimers identify specific inter- and intramolecular interactions and were in good agreement with MS measurements. Specifically, several intermolecular ionic interactions between HER2 Lys-716-HER3 Glu-909, HER2 Glu-717-HER3 Lys-907, and HER2 Asp-871-HER3 Arg-948 were identified by molecular dynamics. We also evaluated the effect of the cancer-associated mutations HER2 D769H/D769Y, HER3 E909G, and HER3 R948K (also numbered HER3 E928G and R967K) on kinase activity in the context of this new structural model. This study provides valuable insights into the EGF receptor/HER/ErbB kinase structure and interactions, which can guide the design of future therapies.
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Carboxyl Group footprinting maps the dimerization interface and phosphorylation induced conformational changes of a membrane associated tyrosine kinase
Molecular & Cellular Proteomics, 2011Co-Authors: Hao Zhang, Michael L Gross, Wei Shen, Don L Rempel, John Monsey, Ilan Vidavsky, Ron BoseAbstract:: Her4 is a transmembrane receptor tyrosine kinase belonging to the ErbB-EGFR family. It plays a vital role in the cardiovascular and nervous systems, and mutations in Her4 have been found in melanoma and lung cancer. The kinase domain of Her4 forms a dimer complex, called the asymmetric dimer, which results in kinase activation. Although a crystal structure of the Her4 asymmetric dimer is known, the dimer affinity and the effect of the subsequent phosphorylation steps on kinase domain conformation are unknown. We report here the use of Carboxyl-Group footprinting MS on a recombinant expressed, Her4 kinase-domain construct to address these questions. Carboxyl-Group footprinting uses a water-soluble carbodiimide, 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide, in the presence of glycine ethyl ester, to modify accessible Carboxyl Groups on glutamate and aspartate residues. Comparisons of Her4 kinase-domain monomers versus dimers and of unphosphorylated versus phosphorylated dimers were made to map the dimerization interface and to determine phosphorylation induced-conformational changes. We detected 37 glutamate and aspartate residues that were modified, and we quantified their extents of modification by liquid chromatography MS. Five residues showed changes in Carboxyl-Group modification. Three of these residues are at the predicted dimer interface, as shown by the crystal structure, and the remaining two residues are on loops that likely have altered conformation in the kinase dimer. Incubating the Her4 kinase dimers with ATP resulted in dramatic increase in Tyr-850 phosphorylation, located on the activation loop, and this resulted in a conformational change in this loop, as evidenced by reduction in Carboxyl-Group modification. The kinase monomer-dimer equilibrium was measured using a titration format in which the extent of Carboxyl-Group footprinting was mathematically modeled to give the dimer association constant (1.5-6.8 × 10(12) dm(2)/mol). This suggests that the kinase-domain makes a significant contribution to the overall dimerization affinity of the full-length Her4 protein.