EPR Studies of V-ATPase with Spin-Labeled Inhibitors DCC and Archazolid: Interaction Dynamics with Proton Translocating Subunit c

Autor(en): Goelz, Jan Philipp 
Bockelmann, Svenja 
Mayer, Kerstin
Steinhoff, Heinz-Juergen 
Wieczorek, Helmut 
Huss, Markus 
Klare, Johann P.
Menche, Dirk 
Stichwörter: ARCHANGIUM-GEPHYRA; archazolid; BIOLOGICAL EVALUATION; BREAST-CANCER; Chemistry, Medicinal; EPR spectroscopy; HIGHLY POTENT; HUMAN PANCREATIC-CANCER; MEDIATED CROSS-LINKING; MYXOBACTERIUM CYSTOBACTER-VIOLACEUS; natural products; Pharmacology & Pharmacy; PLASMA-MEMBRANE; spin labels; STRUCTURAL ELUCIDATION; V-ATPase; VACUOLAR H+-ATPASE
Erscheinungsdatum: 2016
Herausgeber: WILEY-V C H VERLAG GMBH
Enthalten in: CHEMMEDCHEM
Band: 11
Ausgabe: 4
Startseite: 420
Seitenende: 428
Zusammenfassung: 
Vacuolar-type H+-ATPases (V-ATPases) have gained recent attention as highly promising anticancer drug targets, and therefore detailed structural analyses and studies of inhibitor interactions are very important research objectives. Spin labeling of the V-ATPase holoenzyme from the tobacco hornworm Manduca sexta and V-ATPase in isolated yeast (Saccharomyces cerevisiae) vacuoles was accomplished by two novel methods involving the covalent binding of a (2,2,6,6-tetramethylpiperidin-1-yl)oxyl (TEMPO) derivative of N,N-dicyclohexylcarbodiimide (DCC) to the essential glutamate residue in the active site and the noncovalent interaction of a radical analogue of the highly potent inhibitor archazolid, a natural product from myxobacteria. Both complexes were evaluated in detail by electron paramagnetic resonance (EPR) spectroscopic studies and double electron-electron resonance (DEER) measurements, revealing insight into the inhibitor binding mode, dynamics, and stoichiometry as well as into the structure of the central functional subunitc of these medicinally important hetero-multimeric proton-translocating proteins. This study also demonstrates the usefulness of natural product derived spin labels as tools in medicinal chemistry.
ISSN: 18607179
DOI: 10.1002/cmdc.201500500

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