title: Microarray-based screening of heat shock protein inhibitors.
abstract: Based on the importance of heat shock proteins (HSPs) in diseases such as cancer, Alzheimer's disease or malaria, inhibitors of these chaperons are needed. Today's state-of-the-art techniques to identify HSP inhibitors are performed in microplate format, requiring large amounts of proteins and potential inhibitors. In contrast, we have developed a miniaturized protein microarray-based assay to identify novel inhibitors, allowing analysis with 300 pmol of protein. The assay is based on competitive binding of fluorescence-labeled ATP and potential inhibitors to the ATP-binding site of HSP. Therefore, the developed microarray enables the parallel analysis of different ATP-binding proteins on a single microarray. We have demonstrated the possibility of multiplexing by immobilizing full-length human HSP90α and HtpG of Helicobacter pylori on microarrays. Fluorescence-labeled ATP was competed by novel geldanamycin/reblastatin derivatives with IC50 values in the range of 0.5 nM to 4 μM and Z(*)-factors between 0.60 and 0.96. Our results demonstrate the potential of a target-oriented multiplexed protein microarray to identify novel inhibitors for different members of the HSP90 family.
authors.affiliation: Leibniz Universität Hannover, Institut für Technische Chemie and Biomolekulares Wirkstoffzentrum (BMWZ), Callinstr. 5, D-30167 Hannover, Germany.
authors.forename: Emilia
authors.initials: E
authors.lastname: Schax
authors.affiliation: Leibniz Universität Hannover, Institut für Technische Chemie and Biomolekulares Wirkstoffzentrum (BMWZ), Callinstr. 5, D-30167 Hannover, Germany.
authors.forename: Johanna-Gabriela
authors.initials: JG
authors.lastname: Walter
authors.affiliation: Leibniz Universität Hannover, Institut für Technische Chemie and Biomolekulares Wirkstoffzentrum (BMWZ), Callinstr. 5, D-30167 Hannover, Germany.
authors.forename: Helene
authors.initials: H
authors.lastname: Märzhäuser
authors.affiliation: Leibniz Universität Hannover, Institut für Technische Chemie and Biomolekulares Wirkstoffzentrum (BMWZ), Callinstr. 5, D-30167 Hannover, Germany.
authors.forename: Frank
authors.initials: F
authors.lastname: Stahl
authors.affiliation: Leibniz Universität Hannover, Institut für Technische Chemie and Biomolekulares Wirkstoffzentrum (BMWZ), Callinstr. 5, D-30167 Hannover, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Scheper
authors.affiliation: Howard Hughes Medical Institute and the Department of Biochemistry & Biophysics, University of California, San Francisco, 600 16th Street, San Francisco, CA 94158-2517, USA.
authors.forename: David A
authors.initials: DA
authors.lastname: Agard
authors.affiliation: Leibniz Universität Hannover, Institut für Organische Chemie and Biomolekulares Wirkstoffzentrum (BMWZ), Schneiderberg 1, D-30167 Hannover, Germany.
authors.forename: Simone
authors.initials: S
authors.lastname: Eichner
authors.affiliation: Leibniz Universität Hannover, Institut für Organische Chemie and Biomolekulares Wirkstoffzentrum (BMWZ), Schneiderberg 1, D-30167 Hannover, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Kirschning
authors.affiliation: Leibniz Universität Hannover, Institut für Biophysik and Biomolekulares Wirkstoffzentrum (BMWZ), Herrenhäuserstr. 2, D-30167 Hannover, Germany. Electronic address: zeilinger@biophysik.uni-hannover.de.
authors.forename: Carsten
authors.initials: C
authors.lastname: Zeilinger
mesh_terms: D016227:Benzoquinones / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D001667:Binding, Competitive; D004305:Dose-Response Relationship, Drug; D055808:Drug Discovery / Q000379:methods*; D018841:HSP90 Heat-Shock Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D055767:Immobilized Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D047029:Lactams, Macrocyclic / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D040081:Protein Array Analysis / Q000379:methods*; D055550:Protein Stability; D015203:Reproducibility of Results
chemical_list: D016227:Benzoquinones; D018841:HSP90 Heat-Shock Proteins; D055767:Immobilized Proteins; D047029:Lactams, Macrocyclic; C001277:geldanamycin
keywords: Geldanamycin; Heat shock protein; Inhibitor; Protein microarray