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Paul Declerck - One of the best experts on this subject based on the ideXlab platform.

  • importance of the hinge region between α helix f and the main part of serpins based upon identification of the epitope of plasminogen activator inhibitor type 1 neutralizing antibodies
    Journal of Biological Chemistry, 2000
    Co-Authors: A P Bijnens, Ann Gils, I Knockaert, Jan Stassen, Paul Declerck
    Abstract:

    Abstract The serpin plasminogen activator inhibitor type 1 (PAI-1) is an important protein in the regulation of fibrinolysis and inhibits its target proteinases through formation of a covalent complex. In the present study, we have identified the epitope of two PAI-1 neutralizing monoclonal antibodies (MA-33H1F7 and MA-55F4C12). Based upon differential cross-reactivity data of these Monoclonals with PAI-1 from different species and on a sequence alignment between these PAI-1s, combined with the three-dimensional structure, we predicted that the residues Glu128-Val129-Glu130-Arg131and Lys154 (at the hinge region between α-helix F and the main part of the PAI-1-molecule) might form the major site of interaction. Therefore a variety of alanine mutants were generated and evaluated for their affinity toward both monoclonal antibodies. The affinity constants of MA-55F4C12 and MA-33H1F7 for PAI-1 were 2.7 ± 1.6 × 109 m −1 and 5.4 ± 1.7 × 109 m −1, respectively, but decreased between 13- and 270-fold upon mutation of Lys154 to Ala154 or Glu128-Val129-Glu130-Arg131to Ala-Ala-Ala-Ala. The combined mutations (PAI-1-EVER/K), however, resulted in an absence of binding to either of the antibodies. Both antibodies bound to PAI-1-wt/t-PA complexes with a similar affinity as to PAI-1-wt (KA = 4–5 × 109 m −1). The epitope localization reveals the molecular basis for the neutralizing properties of both monoclonal antibodies. In addition, it provides new insights into the validity of various models that have been proposed for the serpin/proteinase complex, excluding full insertion of the reactive-site loop.

  • importance of the hinge region between alpha helix f and the main part of serpins based upon identification of the epitope of plasminogen activator inhibitor type 1 neutralizing antibodies
    Journal of Biological Chemistry, 2000
    Co-Authors: A P Bijnens, Ann Gils, I Knockaert, Jan Stassen, Paul Declerck
    Abstract:

    Abstract The serpin plasminogen activator inhibitor type 1 (PAI-1) is an important protein in the regulation of fibrinolysis and inhibits its target proteinases through formation of a covalent complex. In the present study, we have identified the epitope of two PAI-1 neutralizing monoclonal antibodies (MA-33H1F7 and MA-55F4C12). Based upon differential cross-reactivity data of these Monoclonals with PAI-1 from different species and on a sequence alignment between these PAI-1s, combined with the three-dimensional structure, we predicted that the residues Glu128-Val129-Glu130-Arg131and Lys154 (at the hinge region between α-helix F and the main part of the PAI-1-molecule) might form the major site of interaction. Therefore a variety of alanine mutants were generated and evaluated for their affinity toward both monoclonal antibodies. The affinity constants of MA-55F4C12 and MA-33H1F7 for PAI-1 were 2.7 ± 1.6 × 109 m −1 and 5.4 ± 1.7 × 109 m −1, respectively, but decreased between 13- and 270-fold upon mutation of Lys154 to Ala154 or Glu128-Val129-Glu130-Arg131to Ala-Ala-Ala-Ala. The combined mutations (PAI-1-EVER/K), however, resulted in an absence of binding to either of the antibodies. Both antibodies bound to PAI-1-wt/t-PA complexes with a similar affinity as to PAI-1-wt (KA = 4–5 × 109 m −1). The epitope localization reveals the molecular basis for the neutralizing properties of both monoclonal antibodies. In addition, it provides new insights into the validity of various models that have been proposed for the serpin/proteinase complex, excluding full insertion of the reactive-site loop.

A P Bijnens - One of the best experts on this subject based on the ideXlab platform.

  • importance of the hinge region between α helix f and the main part of serpins based upon identification of the epitope of plasminogen activator inhibitor type 1 neutralizing antibodies
    Journal of Biological Chemistry, 2000
    Co-Authors: A P Bijnens, Ann Gils, I Knockaert, Jan Stassen, Paul Declerck
    Abstract:

    Abstract The serpin plasminogen activator inhibitor type 1 (PAI-1) is an important protein in the regulation of fibrinolysis and inhibits its target proteinases through formation of a covalent complex. In the present study, we have identified the epitope of two PAI-1 neutralizing monoclonal antibodies (MA-33H1F7 and MA-55F4C12). Based upon differential cross-reactivity data of these Monoclonals with PAI-1 from different species and on a sequence alignment between these PAI-1s, combined with the three-dimensional structure, we predicted that the residues Glu128-Val129-Glu130-Arg131and Lys154 (at the hinge region between α-helix F and the main part of the PAI-1-molecule) might form the major site of interaction. Therefore a variety of alanine mutants were generated and evaluated for their affinity toward both monoclonal antibodies. The affinity constants of MA-55F4C12 and MA-33H1F7 for PAI-1 were 2.7 ± 1.6 × 109 m −1 and 5.4 ± 1.7 × 109 m −1, respectively, but decreased between 13- and 270-fold upon mutation of Lys154 to Ala154 or Glu128-Val129-Glu130-Arg131to Ala-Ala-Ala-Ala. The combined mutations (PAI-1-EVER/K), however, resulted in an absence of binding to either of the antibodies. Both antibodies bound to PAI-1-wt/t-PA complexes with a similar affinity as to PAI-1-wt (KA = 4–5 × 109 m −1). The epitope localization reveals the molecular basis for the neutralizing properties of both monoclonal antibodies. In addition, it provides new insights into the validity of various models that have been proposed for the serpin/proteinase complex, excluding full insertion of the reactive-site loop.

  • importance of the hinge region between alpha helix f and the main part of serpins based upon identification of the epitope of plasminogen activator inhibitor type 1 neutralizing antibodies
    Journal of Biological Chemistry, 2000
    Co-Authors: A P Bijnens, Ann Gils, I Knockaert, Jan Stassen, Paul Declerck
    Abstract:

    Abstract The serpin plasminogen activator inhibitor type 1 (PAI-1) is an important protein in the regulation of fibrinolysis and inhibits its target proteinases through formation of a covalent complex. In the present study, we have identified the epitope of two PAI-1 neutralizing monoclonal antibodies (MA-33H1F7 and MA-55F4C12). Based upon differential cross-reactivity data of these Monoclonals with PAI-1 from different species and on a sequence alignment between these PAI-1s, combined with the three-dimensional structure, we predicted that the residues Glu128-Val129-Glu130-Arg131and Lys154 (at the hinge region between α-helix F and the main part of the PAI-1-molecule) might form the major site of interaction. Therefore a variety of alanine mutants were generated and evaluated for their affinity toward both monoclonal antibodies. The affinity constants of MA-55F4C12 and MA-33H1F7 for PAI-1 were 2.7 ± 1.6 × 109 m −1 and 5.4 ± 1.7 × 109 m −1, respectively, but decreased between 13- and 270-fold upon mutation of Lys154 to Ala154 or Glu128-Val129-Glu130-Arg131to Ala-Ala-Ala-Ala. The combined mutations (PAI-1-EVER/K), however, resulted in an absence of binding to either of the antibodies. Both antibodies bound to PAI-1-wt/t-PA complexes with a similar affinity as to PAI-1-wt (KA = 4–5 × 109 m −1). The epitope localization reveals the molecular basis for the neutralizing properties of both monoclonal antibodies. In addition, it provides new insights into the validity of various models that have been proposed for the serpin/proteinase complex, excluding full insertion of the reactive-site loop.

Jan Stassen - One of the best experts on this subject based on the ideXlab platform.

  • importance of the hinge region between α helix f and the main part of serpins based upon identification of the epitope of plasminogen activator inhibitor type 1 neutralizing antibodies
    Journal of Biological Chemistry, 2000
    Co-Authors: A P Bijnens, Ann Gils, I Knockaert, Jan Stassen, Paul Declerck
    Abstract:

    Abstract The serpin plasminogen activator inhibitor type 1 (PAI-1) is an important protein in the regulation of fibrinolysis and inhibits its target proteinases through formation of a covalent complex. In the present study, we have identified the epitope of two PAI-1 neutralizing monoclonal antibodies (MA-33H1F7 and MA-55F4C12). Based upon differential cross-reactivity data of these Monoclonals with PAI-1 from different species and on a sequence alignment between these PAI-1s, combined with the three-dimensional structure, we predicted that the residues Glu128-Val129-Glu130-Arg131and Lys154 (at the hinge region between α-helix F and the main part of the PAI-1-molecule) might form the major site of interaction. Therefore a variety of alanine mutants were generated and evaluated for their affinity toward both monoclonal antibodies. The affinity constants of MA-55F4C12 and MA-33H1F7 for PAI-1 were 2.7 ± 1.6 × 109 m −1 and 5.4 ± 1.7 × 109 m −1, respectively, but decreased between 13- and 270-fold upon mutation of Lys154 to Ala154 or Glu128-Val129-Glu130-Arg131to Ala-Ala-Ala-Ala. The combined mutations (PAI-1-EVER/K), however, resulted in an absence of binding to either of the antibodies. Both antibodies bound to PAI-1-wt/t-PA complexes with a similar affinity as to PAI-1-wt (KA = 4–5 × 109 m −1). The epitope localization reveals the molecular basis for the neutralizing properties of both monoclonal antibodies. In addition, it provides new insights into the validity of various models that have been proposed for the serpin/proteinase complex, excluding full insertion of the reactive-site loop.

  • importance of the hinge region between alpha helix f and the main part of serpins based upon identification of the epitope of plasminogen activator inhibitor type 1 neutralizing antibodies
    Journal of Biological Chemistry, 2000
    Co-Authors: A P Bijnens, Ann Gils, I Knockaert, Jan Stassen, Paul Declerck
    Abstract:

    Abstract The serpin plasminogen activator inhibitor type 1 (PAI-1) is an important protein in the regulation of fibrinolysis and inhibits its target proteinases through formation of a covalent complex. In the present study, we have identified the epitope of two PAI-1 neutralizing monoclonal antibodies (MA-33H1F7 and MA-55F4C12). Based upon differential cross-reactivity data of these Monoclonals with PAI-1 from different species and on a sequence alignment between these PAI-1s, combined with the three-dimensional structure, we predicted that the residues Glu128-Val129-Glu130-Arg131and Lys154 (at the hinge region between α-helix F and the main part of the PAI-1-molecule) might form the major site of interaction. Therefore a variety of alanine mutants were generated and evaluated for their affinity toward both monoclonal antibodies. The affinity constants of MA-55F4C12 and MA-33H1F7 for PAI-1 were 2.7 ± 1.6 × 109 m −1 and 5.4 ± 1.7 × 109 m −1, respectively, but decreased between 13- and 270-fold upon mutation of Lys154 to Ala154 or Glu128-Val129-Glu130-Arg131to Ala-Ala-Ala-Ala. The combined mutations (PAI-1-EVER/K), however, resulted in an absence of binding to either of the antibodies. Both antibodies bound to PAI-1-wt/t-PA complexes with a similar affinity as to PAI-1-wt (KA = 4–5 × 109 m −1). The epitope localization reveals the molecular basis for the neutralizing properties of both monoclonal antibodies. In addition, it provides new insights into the validity of various models that have been proposed for the serpin/proteinase complex, excluding full insertion of the reactive-site loop.

I Knockaert - One of the best experts on this subject based on the ideXlab platform.

  • importance of the hinge region between α helix f and the main part of serpins based upon identification of the epitope of plasminogen activator inhibitor type 1 neutralizing antibodies
    Journal of Biological Chemistry, 2000
    Co-Authors: A P Bijnens, Ann Gils, I Knockaert, Jan Stassen, Paul Declerck
    Abstract:

    Abstract The serpin plasminogen activator inhibitor type 1 (PAI-1) is an important protein in the regulation of fibrinolysis and inhibits its target proteinases through formation of a covalent complex. In the present study, we have identified the epitope of two PAI-1 neutralizing monoclonal antibodies (MA-33H1F7 and MA-55F4C12). Based upon differential cross-reactivity data of these Monoclonals with PAI-1 from different species and on a sequence alignment between these PAI-1s, combined with the three-dimensional structure, we predicted that the residues Glu128-Val129-Glu130-Arg131and Lys154 (at the hinge region between α-helix F and the main part of the PAI-1-molecule) might form the major site of interaction. Therefore a variety of alanine mutants were generated and evaluated for their affinity toward both monoclonal antibodies. The affinity constants of MA-55F4C12 and MA-33H1F7 for PAI-1 were 2.7 ± 1.6 × 109 m −1 and 5.4 ± 1.7 × 109 m −1, respectively, but decreased between 13- and 270-fold upon mutation of Lys154 to Ala154 or Glu128-Val129-Glu130-Arg131to Ala-Ala-Ala-Ala. The combined mutations (PAI-1-EVER/K), however, resulted in an absence of binding to either of the antibodies. Both antibodies bound to PAI-1-wt/t-PA complexes with a similar affinity as to PAI-1-wt (KA = 4–5 × 109 m −1). The epitope localization reveals the molecular basis for the neutralizing properties of both monoclonal antibodies. In addition, it provides new insights into the validity of various models that have been proposed for the serpin/proteinase complex, excluding full insertion of the reactive-site loop.

  • importance of the hinge region between alpha helix f and the main part of serpins based upon identification of the epitope of plasminogen activator inhibitor type 1 neutralizing antibodies
    Journal of Biological Chemistry, 2000
    Co-Authors: A P Bijnens, Ann Gils, I Knockaert, Jan Stassen, Paul Declerck
    Abstract:

    Abstract The serpin plasminogen activator inhibitor type 1 (PAI-1) is an important protein in the regulation of fibrinolysis and inhibits its target proteinases through formation of a covalent complex. In the present study, we have identified the epitope of two PAI-1 neutralizing monoclonal antibodies (MA-33H1F7 and MA-55F4C12). Based upon differential cross-reactivity data of these Monoclonals with PAI-1 from different species and on a sequence alignment between these PAI-1s, combined with the three-dimensional structure, we predicted that the residues Glu128-Val129-Glu130-Arg131and Lys154 (at the hinge region between α-helix F and the main part of the PAI-1-molecule) might form the major site of interaction. Therefore a variety of alanine mutants were generated and evaluated for their affinity toward both monoclonal antibodies. The affinity constants of MA-55F4C12 and MA-33H1F7 for PAI-1 were 2.7 ± 1.6 × 109 m −1 and 5.4 ± 1.7 × 109 m −1, respectively, but decreased between 13- and 270-fold upon mutation of Lys154 to Ala154 or Glu128-Val129-Glu130-Arg131to Ala-Ala-Ala-Ala. The combined mutations (PAI-1-EVER/K), however, resulted in an absence of binding to either of the antibodies. Both antibodies bound to PAI-1-wt/t-PA complexes with a similar affinity as to PAI-1-wt (KA = 4–5 × 109 m −1). The epitope localization reveals the molecular basis for the neutralizing properties of both monoclonal antibodies. In addition, it provides new insights into the validity of various models that have been proposed for the serpin/proteinase complex, excluding full insertion of the reactive-site loop.

Ann Gils - One of the best experts on this subject based on the ideXlab platform.

  • importance of the hinge region between α helix f and the main part of serpins based upon identification of the epitope of plasminogen activator inhibitor type 1 neutralizing antibodies
    Journal of Biological Chemistry, 2000
    Co-Authors: A P Bijnens, Ann Gils, I Knockaert, Jan Stassen, Paul Declerck
    Abstract:

    Abstract The serpin plasminogen activator inhibitor type 1 (PAI-1) is an important protein in the regulation of fibrinolysis and inhibits its target proteinases through formation of a covalent complex. In the present study, we have identified the epitope of two PAI-1 neutralizing monoclonal antibodies (MA-33H1F7 and MA-55F4C12). Based upon differential cross-reactivity data of these Monoclonals with PAI-1 from different species and on a sequence alignment between these PAI-1s, combined with the three-dimensional structure, we predicted that the residues Glu128-Val129-Glu130-Arg131and Lys154 (at the hinge region between α-helix F and the main part of the PAI-1-molecule) might form the major site of interaction. Therefore a variety of alanine mutants were generated and evaluated for their affinity toward both monoclonal antibodies. The affinity constants of MA-55F4C12 and MA-33H1F7 for PAI-1 were 2.7 ± 1.6 × 109 m −1 and 5.4 ± 1.7 × 109 m −1, respectively, but decreased between 13- and 270-fold upon mutation of Lys154 to Ala154 or Glu128-Val129-Glu130-Arg131to Ala-Ala-Ala-Ala. The combined mutations (PAI-1-EVER/K), however, resulted in an absence of binding to either of the antibodies. Both antibodies bound to PAI-1-wt/t-PA complexes with a similar affinity as to PAI-1-wt (KA = 4–5 × 109 m −1). The epitope localization reveals the molecular basis for the neutralizing properties of both monoclonal antibodies. In addition, it provides new insights into the validity of various models that have been proposed for the serpin/proteinase complex, excluding full insertion of the reactive-site loop.

  • importance of the hinge region between alpha helix f and the main part of serpins based upon identification of the epitope of plasminogen activator inhibitor type 1 neutralizing antibodies
    Journal of Biological Chemistry, 2000
    Co-Authors: A P Bijnens, Ann Gils, I Knockaert, Jan Stassen, Paul Declerck
    Abstract:

    Abstract The serpin plasminogen activator inhibitor type 1 (PAI-1) is an important protein in the regulation of fibrinolysis and inhibits its target proteinases through formation of a covalent complex. In the present study, we have identified the epitope of two PAI-1 neutralizing monoclonal antibodies (MA-33H1F7 and MA-55F4C12). Based upon differential cross-reactivity data of these Monoclonals with PAI-1 from different species and on a sequence alignment between these PAI-1s, combined with the three-dimensional structure, we predicted that the residues Glu128-Val129-Glu130-Arg131and Lys154 (at the hinge region between α-helix F and the main part of the PAI-1-molecule) might form the major site of interaction. Therefore a variety of alanine mutants were generated and evaluated for their affinity toward both monoclonal antibodies. The affinity constants of MA-55F4C12 and MA-33H1F7 for PAI-1 were 2.7 ± 1.6 × 109 m −1 and 5.4 ± 1.7 × 109 m −1, respectively, but decreased between 13- and 270-fold upon mutation of Lys154 to Ala154 or Glu128-Val129-Glu130-Arg131to Ala-Ala-Ala-Ala. The combined mutations (PAI-1-EVER/K), however, resulted in an absence of binding to either of the antibodies. Both antibodies bound to PAI-1-wt/t-PA complexes with a similar affinity as to PAI-1-wt (KA = 4–5 × 109 m −1). The epitope localization reveals the molecular basis for the neutralizing properties of both monoclonal antibodies. In addition, it provides new insights into the validity of various models that have been proposed for the serpin/proteinase complex, excluding full insertion of the reactive-site loop.