title: Chemical screens against a reconstituted multiprotein complex: myricetin blocks DnaJ regulation of DnaK through an allosteric mechanism.
abstract: DnaK is a molecular chaperone responsible for multiple aspects of bacterial proteostasis. The intrinsically slow ATPase activity of DnaK is stimulated by its co-chaperone, DnaJ, and these proteins often work in concert. To identify inhibitors we screened plant-derived extracts against a reconstituted mixture of DnaK and DnaJ. This approach resulted in the identification of flavonoids, including myricetin, which inhibited activity by up to 75%. Interestingly, myricetin prevented DnaJ-mediated stimulation of ATPase activity, with minimal impact on either DnaK's intrinsic turnover rate or its stimulation by another co-chaperone, GrpE. Using NMR, we found that myricetin binds DnaK at an unanticipated site between the IB and IIB subdomains and that it allosterically blocked binding of DnaK to DnaJ. Together, these results highlight a "gray box" screening approach, which might facilitate the identification of inhibitors of other protein-protein interactions.
authors.affiliation: Chemical Biology Graduate Program, University of Michigan, Ann Arbor, MI 48109, USA.
authors.forename: Lyra
authors.initials: L
authors.lastname: Chang
authors.forename: Yoshinari
authors.initials: Y
authors.lastname: Miyata
authors.forename: Peter M U
authors.initials: PM
authors.lastname: Ung
authors.forename: Eric B
authors.initials: EB
authors.lastname: Bertelsen
authors.forename: Thomas J
authors.initials: TJ
authors.lastname: McQuade
authors.forename: Heather A
authors.initials: HA
authors.lastname: Carlson
authors.forename: Erik R P
authors.initials: ER
authors.lastname: Zuiderweg
authors.forename: Jason E
authors.initials: JE
authors.lastname: Gestwicki
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000494:Allosteric Regulation / Q000187:drug effects; D004353:Drug Evaluation, Preclinical; D029968:Escherichia coli Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D005419:Flavonoids / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D050956:HSP40 Heat-Shock Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D018840:HSP70 Heat-Shock Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D008958:Models, Molecular; D010936:Plant Extracts / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D017434:Protein Structure, Tertiary; D013329:Structure-Activity Relationship
chemical_list: D029968:Escherichia coli Proteins; D005419:Flavonoids; D050956:HSP40 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins; D010936:Plant Extracts; C040015:myricetin; D000255:Adenosine Triphosphate; C063842:dnaK protein, E coli