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Florian Stengel - One of the best experts on this subject based on the ideXlab platform.

  • structural dynamics of the e6ap ube3a e6 p53 enzyme Substrate Complex
    Nature Communications, 2018
    Co-Authors: Carolin Sailer, Fabian Offensperger, Alexandra Julier, Kaimichael Kammer, Ryan Walkergray, Matthew G Gold, Martin Scheffner, Florian Stengel
    Abstract:

    Deregulation of the ubiquitin ligase E6AP is causally linked to the development of human disease, including cervical cancer. In Complex with the E6 oncoprotein of human papillomaviruses, E6AP targets the tumor suppressor p53 for degradation, thereby contributing to carcinogenesis. Moreover, E6 acts as a potent activator of E6AP by a yet unknown mechanism. However, structural information explaining how the E6AP-E6-p53 enzyme-Substrate Complex is assembled, and how E6 stimulates E6AP, is largely missing. Here, we develop and apply different crosslinking mass spectrometry-based approaches to study the E6AP-E6-p53 interplay. We show that binding of E6 induces conformational rearrangements in E6AP, thereby positioning E6 and p53 in the immediate vicinity of the catalytic center of E6AP. Our data provide structural and functional insights into the dynamics of the full-length E6AP-E6-p53 enzyme-Substrate Complex, demonstrating how E6 can stimulate the ubiquitin ligase activity of E6AP while facilitating ubiquitin transfer from E6AP onto p53.

  • Structural dynamics of the E6AP/UBE3A-E6-p53 enzyme-Substrate Complex.
    Nature Communications, 2018
    Co-Authors: Carolin Sailer, Fabian Offensperger, Alexandra Julier, Kaimichael Kammer, Ryan Walker-gray, Matthew G Gold, Martin Scheffner, Florian Stengel
    Abstract:

    Deregulation of the ubiquitin ligase E6AP is causally linked to the development of human disease, including cervical cancer. In Complex with the E6 oncoprotein of human papillomaviruses, E6AP targets the tumor suppressor p53 for degradation, thereby contributing to carcinogenesis. Moreover, E6 acts as a potent activator of E6AP by a yet unknown mechanism. However, structural information explaining how the E6AP-E6-p53 enzyme-Substrate Complex is assembled, and how E6 stimulates E6AP, is largely missing. Here, we develop and apply different crosslinking mass spectrometry-based approaches to study the E6AP-E6-p53 interplay. We show that binding of E6 induces conformational rearrangements in E6AP, thereby positioning E6 and p53 in the immediate vicinity of the catalytic center of E6AP. Our data provide structural and functional insights into the dynamics of the full-length E6AP-E6-p53 enzyme-Substrate Complex, demonstrating how E6 can stimulate the ubiquitin ligase activity of E6AP while facilitating ubiquitin transfer from E6AP onto p53.

Yuriy F. Zuev - One of the best experts on this subject based on the ideXlab platform.

  • Brownian dynamics simulation of Substrate motion near active site of enzyme entrapped inside reverse micelle.
    European Biophysics Journal, 2010
    Co-Authors: Elena A Ermakova, Natalia L Zakhartchenko, Yuriy F. Zuev
    Abstract:

    Brownian dynamics simulation has been applied to analyze the influence of the electrostatic field of a reverse micelle on the enzyme-Substrate Complex formation inside a micelle. The probability that the enzyme-Substrate Complex will form from serine protease (trypsin) and the specific hydrophilic cationic Substrate Nα-benzoyl-l-arginine ethyl ester has been studied within the framework of the encounter Complex formation theory. It has been shown that surfactant charge, dipole moments created by charged surfactant molecules and counterions, and permittivity of the inner core of reverse micelles can all be used as regulatory parameters to alter the Substrate orientation near the active site of the enzyme and to change the probability that the enzyme-Substrate Complex will form.

  • effect of surface potential of reverse micelle on enzyme Substrate Complex formation
    Colloids and Surfaces A: Physicochemical and Engineering Aspects, 2008
    Co-Authors: Elena A Ermakova, Natalia L Zakhartchenko, Yuriy F. Zuev
    Abstract:

    Abstract The influence of the electrostatic potential of reverse micelle on the enzyme–Substrate Complex formation have been studied within the framework of the encounter Complex (EC) formation theory. Reverse micelles have a multiple-factor effect on encapsulated substances and on mechanisms of their interaction. The action of individual constituents on the overall micellar effect was analyzed by means of Brownian dynamics simulation. The effects of surface potential in charged and neutral reverse micelles, of the size and of the ionic strength of micellar core on the probability to form the reaction Complex between serine protease (trypsin) and specific Substrate Nα-benzoyl- l -arginine ethyl ester was examined. It was shown that negative potential of micelle increases the probability of EC formation and the positive potential decreases it. Orientation of Substrate in electrostatic field of micelle depends on the value and the sign of surface potential.

  • Effect of surface potential of reverse micelle on enzyme–Substrate Complex formation
    Colloids and Surfaces A: Physicochemical and Engineering Aspects, 2008
    Co-Authors: Elena A Ermakova, Natalia L Zakhartchenko, Yuriy F. Zuev
    Abstract:

    Abstract The influence of the electrostatic potential of reverse micelle on the enzyme–Substrate Complex formation have been studied within the framework of the encounter Complex (EC) formation theory. Reverse micelles have a multiple-factor effect on encapsulated substances and on mechanisms of their interaction. The action of individual constituents on the overall micellar effect was analyzed by means of Brownian dynamics simulation. The effects of surface potential in charged and neutral reverse micelles, of the size and of the ionic strength of micellar core on the probability to form the reaction Complex between serine protease (trypsin) and specific Substrate Nα-benzoyl- l -arginine ethyl ester was examined. It was shown that negative potential of micelle increases the probability of EC formation and the positive potential decreases it. Orientation of Substrate in electrostatic field of micelle depends on the value and the sign of surface potential.

Carolin Sailer - One of the best experts on this subject based on the ideXlab platform.

  • structural dynamics of the e6ap ube3a e6 p53 enzyme Substrate Complex
    Nature Communications, 2018
    Co-Authors: Carolin Sailer, Fabian Offensperger, Alexandra Julier, Kaimichael Kammer, Ryan Walkergray, Matthew G Gold, Martin Scheffner, Florian Stengel
    Abstract:

    Deregulation of the ubiquitin ligase E6AP is causally linked to the development of human disease, including cervical cancer. In Complex with the E6 oncoprotein of human papillomaviruses, E6AP targets the tumor suppressor p53 for degradation, thereby contributing to carcinogenesis. Moreover, E6 acts as a potent activator of E6AP by a yet unknown mechanism. However, structural information explaining how the E6AP-E6-p53 enzyme-Substrate Complex is assembled, and how E6 stimulates E6AP, is largely missing. Here, we develop and apply different crosslinking mass spectrometry-based approaches to study the E6AP-E6-p53 interplay. We show that binding of E6 induces conformational rearrangements in E6AP, thereby positioning E6 and p53 in the immediate vicinity of the catalytic center of E6AP. Our data provide structural and functional insights into the dynamics of the full-length E6AP-E6-p53 enzyme-Substrate Complex, demonstrating how E6 can stimulate the ubiquitin ligase activity of E6AP while facilitating ubiquitin transfer from E6AP onto p53.

  • Structural dynamics of the E6AP/UBE3A-E6-p53 enzyme-Substrate Complex.
    Nature Communications, 2018
    Co-Authors: Carolin Sailer, Fabian Offensperger, Alexandra Julier, Kaimichael Kammer, Ryan Walker-gray, Matthew G Gold, Martin Scheffner, Florian Stengel
    Abstract:

    Deregulation of the ubiquitin ligase E6AP is causally linked to the development of human disease, including cervical cancer. In Complex with the E6 oncoprotein of human papillomaviruses, E6AP targets the tumor suppressor p53 for degradation, thereby contributing to carcinogenesis. Moreover, E6 acts as a potent activator of E6AP by a yet unknown mechanism. However, structural information explaining how the E6AP-E6-p53 enzyme-Substrate Complex is assembled, and how E6 stimulates E6AP, is largely missing. Here, we develop and apply different crosslinking mass spectrometry-based approaches to study the E6AP-E6-p53 interplay. We show that binding of E6 induces conformational rearrangements in E6AP, thereby positioning E6 and p53 in the immediate vicinity of the catalytic center of E6AP. Our data provide structural and functional insights into the dynamics of the full-length E6AP-E6-p53 enzyme-Substrate Complex, demonstrating how E6 can stimulate the ubiquitin ligase activity of E6AP while facilitating ubiquitin transfer from E6AP onto p53.

Jeffrey A. Haas - One of the best experts on this subject based on the ideXlab platform.

  • Fluorescence anisotropy studies of enzyme-Substrate Complex formation in stearoyl-ACP desaturase.
    Biochemistry, 2002
    Co-Authors: Jeffrey A. Haas
    Abstract:

    : Stearoyl-acyl carrier protein Delta(9)-desaturase (delta9D) catalyzes regio- and stereospecific insertion of cis double bonds into acyl chains attached to acyl carrier protein. Steady-state and stopped-flow fluorescence anisotropy measurements using acylated forms of dansyl- and fluoresceinyl-ACPs revealed equilibrium dissociation constants and dissociation rate constants for 16:0-, 17:0-, and 18:0-ACPs with resting and chemically 4e(-) reduced delta9D. Binding of 1 nM 18:0-fluoresceinyl-ACP to one subunit of the dimeric resting delta9D was observed with K(D1) = 13 +/- 3 nM. No significant difference in the K(D1) value was observed for 4e(-) delta9D. An approximately 4-fold increase in K(D1) per methylene group was observed upon shortening the acyl chain from 18:0 to 17:0 and then 16:0. In different experiments performed with 850 nM 18:0-dansyl-ACP, binding to the second subunit of resting delta9D was estimated to have K(D2) approximately 350 +/- 40 nM. The K(D2) values exhibited a similar dependence on acyl chain length as observed for the K(D1) values. The k(off) values measured by stopped-flow anisotropy measurements for reversal of the enzyme-Substrate Complex were also acyl-chain length dependent and increased 130-fold for 16:0-ACP (130 s(-)(1)) relative to 18:0-ACP (1 s(-)(1)). Increases in acyl chain length are thus associated with the presently reported increases in the K(D) and k(off) values. These results indicate that acyl chain length selectivity derives in major part from partition of the enzyme-Substrate Complex between Substrate release and subsequent steps in catalysis.

  • fluorescence anisotropy studies of enzyme Substrate Complex formation in stearoyl acp desaturase
    Biochemistry, 2002
    Co-Authors: Jeffrey A. Haas
    Abstract:

    Stearoyl-acyl carrier protein Δ 9-desaturase (Δ9D) catalyzes regio- and stereospecific insertion of cis double bonds into acyl chains attached to acyl carrier protein. Steady-state and stopped-flow...

Elena A Ermakova - One of the best experts on this subject based on the ideXlab platform.

  • Brownian dynamics simulation of Substrate motion near active site of enzyme entrapped inside reverse micelle.
    European Biophysics Journal, 2010
    Co-Authors: Elena A Ermakova, Natalia L Zakhartchenko, Yuriy F. Zuev
    Abstract:

    Brownian dynamics simulation has been applied to analyze the influence of the electrostatic field of a reverse micelle on the enzyme-Substrate Complex formation inside a micelle. The probability that the enzyme-Substrate Complex will form from serine protease (trypsin) and the specific hydrophilic cationic Substrate Nα-benzoyl-l-arginine ethyl ester has been studied within the framework of the encounter Complex formation theory. It has been shown that surfactant charge, dipole moments created by charged surfactant molecules and counterions, and permittivity of the inner core of reverse micelles can all be used as regulatory parameters to alter the Substrate orientation near the active site of the enzyme and to change the probability that the enzyme-Substrate Complex will form.

  • effect of surface potential of reverse micelle on enzyme Substrate Complex formation
    Colloids and Surfaces A: Physicochemical and Engineering Aspects, 2008
    Co-Authors: Elena A Ermakova, Natalia L Zakhartchenko, Yuriy F. Zuev
    Abstract:

    Abstract The influence of the electrostatic potential of reverse micelle on the enzyme–Substrate Complex formation have been studied within the framework of the encounter Complex (EC) formation theory. Reverse micelles have a multiple-factor effect on encapsulated substances and on mechanisms of their interaction. The action of individual constituents on the overall micellar effect was analyzed by means of Brownian dynamics simulation. The effects of surface potential in charged and neutral reverse micelles, of the size and of the ionic strength of micellar core on the probability to form the reaction Complex between serine protease (trypsin) and specific Substrate Nα-benzoyl- l -arginine ethyl ester was examined. It was shown that negative potential of micelle increases the probability of EC formation and the positive potential decreases it. Orientation of Substrate in electrostatic field of micelle depends on the value and the sign of surface potential.

  • Effect of surface potential of reverse micelle on enzyme–Substrate Complex formation
    Colloids and Surfaces A: Physicochemical and Engineering Aspects, 2008
    Co-Authors: Elena A Ermakova, Natalia L Zakhartchenko, Yuriy F. Zuev
    Abstract:

    Abstract The influence of the electrostatic potential of reverse micelle on the enzyme–Substrate Complex formation have been studied within the framework of the encounter Complex (EC) formation theory. Reverse micelles have a multiple-factor effect on encapsulated substances and on mechanisms of their interaction. The action of individual constituents on the overall micellar effect was analyzed by means of Brownian dynamics simulation. The effects of surface potential in charged and neutral reverse micelles, of the size and of the ionic strength of micellar core on the probability to form the reaction Complex between serine protease (trypsin) and specific Substrate Nα-benzoyl- l -arginine ethyl ester was examined. It was shown that negative potential of micelle increases the probability of EC formation and the positive potential decreases it. Orientation of Substrate in electrostatic field of micelle depends on the value and the sign of surface potential.