Linked Life Data

21067198

Source:http://linkedlifedata.com/resource/pubmed/id/21067198

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
23
pubmed:dateCreated
2010-12-1
pubmed:abstractText
Protein-protein interactions are an intricate part of biological pathways and have become important targets for drug discovery. Here we present a two-stage magnetic bead assay to functionally screen small-molecule mixtures for modulators of protein-based interactions, with simultaneous affinity-based isolation of active compounds and identification by mass spectrometry. Proteins of interest interact in solution prior to the addition of Ni(II)-functionalized magnetic beads to recover an intact protein-protein complex through affinity capture of a polyhistidine-tagged primary target ("protein-complex fishing"). Protein-complex fishing, utilizing His(6)-tagged calmodulin (CaM) as the primary (bait) protein and melittin (Mel) as the target, was used to screen a mass-encoded library of 1000 bioactive compounds (50 mixtures, 20 compounds each) and successfully identified three known antagonists, three naturally occurring phenolic compounds previously reported to disrupt CaM-activated phosphodiesterase activity, and two newly identified modulators of the CaM-Mel interaction, methylbenzethonium and pempidine tartrate. The ability to produce quantitative inhibition data is also shown through the development of dose-dependent response curves and the determination of inhibition constants (K(I)) for the novel compound methylbenzethonium (K(I) = 14-49 nM) and two known antagonists, calmidazolium (K(I) = 1.7-7.5 nM) and trifluoperazine (K(I) = 1.2-3.0 ?M), with the latter two values being in close agreement with literature values.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Dec
pubmed:issn
1520-6882
pubmed:author
pubmed:issnType
Electronic
pubmed:day
1
pubmed:volume
82
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
9850-7
pubmed:meshHeading
pubmed:year
2010
pubmed:articleTitle
Magnetic "fishing" assay to screen small-molecule mixtures for modulators of protein-protein interactions.
pubmed:affiliation
Chemical Biology Graduate Program, Department of Chemistry and Chemical Biology, McMaster University, 1280 Main Street West, Hamilton, Ontario L8S 4M1, Canada.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't