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ABSTRACT: Libraries of small molecules were searched for Fc-fragment selective binders to a recombinant human antibody ("MDJ8″, IgG(1)-subtype, κ-light chain) via SPR-based screening of chemical microarrays. Identified hit structures were immobilised on NHS-activated Sepharose for the determination of MDJ8 binding and selectivity versus typical proteineous impurities represented by the spend cell culture supernatant. Columns were packed and the most promising ligands further characterized in terms of binding constants, binding kinetics, as well as dynamic and equilibrium binding capacities. The performance of the best ligand, 2A10, was compared to standard Protein A chromatography. Using ligand 2A10 antibody capture from unprocessed cell culture supernatants was possible at similar recovery yield (>90%), purity (>80%), and eluting concentration (approximately 1 g/L) as with Protein A. Affinity constants (K(d)) of 2A10 were an order of magnitude higher than for the Protein A material, but still in the nM-range, while maximum binding capacities and binding kinetics were in the same order of magnitude. Ligand 2A10 was also able to capture a murine monoclonal antibody, again with similar efficiency as Protein A, as well as a number of humanised therapeutic antibodies. Antibody elution from the 2A10 column was possible using the Protein A standard protocol, i.e. 100mM glycine HCl pH 3.0, but also at near physiological pH, when some organic solvent or modifiers were present. Ligand 2A10 thus constitutes a cheaper, more robust alternative to Protein A as possible generic antibody binder. Moreover, the outlined approach to ligand selection could in principle by used to create suitable affinity ligands for other high value biotech products.
Journal of chromatography. A 07/2011; 1218(29):4649-59. · 4.19 Impact Factor
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Alexander Heim-Riether,
Steven J Taylor,
Shuang Liang,
Donghong Amy Gao,
Zhaoming Xiong,
E Michael August,
Brandon K Collins,
Bennett T Farmer,
Kathleen Haverty,
Melissa Hill-Drzewi,
Hans-Dieter Junker,
S Mariana Margarit,
Neil Moss,
Thomas Neumann,
John R Proudfoot,
Lana Smith Keenan, Renate Sekul,
Qiang Zhang,
Jun Li,
Neil A Farrow
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ABSTRACT: Discovery and optimization of potency and selectivity of a non-Zn-chelating MMP-13 inhibitor with the aid of protein co-crystal structural information is reported. This inhibitor was observed to have a binding mode distinct from previously published MMP-13 inhibitors. Potency and selectivity were improved by extending the hit structure out from the active site into the S1' pocket.
Bioorganic & medicinal chemistry letters 09/2009; 19(18):5321-4. · 2.65 Impact Factor
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Thomas Neumann,
Hans-Dieter Junker,
Oliver Keil,
Klaus Burkert,
Holger Ottleben,
Jurgen Gamer, Renate Sekul,
Holger Deppe,
Achim Feurer,
Dirk Tomandl,
Gunther Metz
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ABSTRACT: Surface plasmon resonance imaging, a low affinity screening method, allows the highly parallel detection of small molecules binding to a target protein. The screening of a fragment based compound library immobilized on chemical microarrays resulted in the discovery of binding fragments for the serine protease thrombin. Functional assays confirmed enzymatic inhibition of microarray hits and crystallography established the binding mode of a non-basic S1 motif providing a starting point for medicinal chemistry.
Letters in Drug Design & Discovery 11/2005; 2(8):590-594. · 0.87 Impact Factor
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Stefan Dickopf,
Michael Frank,
Hans-Dieter Junker,
Sabine Maier,
Günther Metz,
Holger Ottleben,
Harald Rau,
Nathalie Schellhaas,
Kristina Schmidt, Renate Sekul,
Cecile Vanier,
Dirk Vetter,
Jörg Czech,
Martin Lorenz,
Hans Matter,
Manfred Schudok,
Herman Schreuder,
David W Will,
Hans Peter Nestler
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ABSTRACT: The goal of this study was to explore the applicability of surface plasmon resonance (SPR)-based fragment screening to identify compounds that bind to factor VIIa (FVIIa). Based on pharmacophore models virtual screening approaches, we selected fragments anticipated to have a reasonable chance of binding to the S1-binding pocket of FVIIa and immobilized these compounds on microarrays. In affinity fingerprinting experiments, a number of compounds were identified to be specifically interacting with FVIIa and shown to fall into four structural classes. The results demonstrate that the chemical microarray technology platform using SPR detection generates unique chemobiological information that is useful for de novo discovery and lead development and allows the detection of weak interactions with ligands of low molecular weight.
Analytical Biochemistry 01/2005; 335(1):50-7. · 3.00 Impact Factor
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Alexander Heim-Riether,
Steven J. Taylor,
Shuang Liang,
Donghong Amy Gao,
Zhaoming Xiong,
E. Michael August,
Brandon K. Collins,
Bennett T. Farmer II,
Kathleen Haverty,
Melissa Hill-Drzewi,
Hans-Dieter Junker,
S. Mariana Margarit,
Neil Moss,
Thomas Neumann,
John R. Proudfoot,
Lana Smith Keenan, Renate Sekul,
Qiang Zhang,
Jun Li,
Neil A. Farrow
Bioorganic & Medicinal Chemistry Letters. 19(18):5321-5324.
