A three-hybrid approach to scanning the proteome for targets of small molecule kinase inhibitors.
In this study, we explored the application of a yeast three-hybrid (Y3H)-based compound/protein display system to scanning the proteome for targets of kinase inhibitors. Various known cyclin-dependent kinase (CDK) inhibitors, including purine and indenopyrazole analogs, were displayed in the form of methotrexate-based hybrid ligands and deployed in cDNA library or yeast cell array-based screening formats. For all inhibitors, known cell cycle CDKs as well as novel candidate CDK-like and/or CDK-unrelated kinase targets could be identified, many of which were independently confirmed using secondary enzyme assays and affinity chromatography. The Y3H system described here may prove generally useful in the discovery of candidate drug targets.
Becker
Frank
F
GPC Biotech AG, 20 Fraunhoferstrasse, Planegg/Martinsried 82152, Germany.
Murthi
Krishna
K
Smith
Chase
C
Come
Jon
J
Costa-Roldán
Nuria
N
Kaufmann
Christine
C
Hanke
Urs
U
Degenhart
Carsten
C
Baumann
Sabine
S
Wallner
Wolfgang
W
Huber
Andrea
A
Dedier
Severine
S
Dill
Simone
S
Kinsman
David
D
Hediger
Mark
M
Bockovich
Nicholas
N
Meier-Ewert
Sebastian
S
Kluge
Arthur F
AF
Kley
Nikolai
N
D000225:Adenine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D016203:CDC2 Protein Kinase / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism; D018844:Cyclin-Dependent Kinases / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D008024:Ligands; D040081:Protein Array Analysis; D020543:Proteome / Q000235:genetics; D012440:Saccharomyces / Q000235:genetics; D020798:Two-Hybrid System Techniques
D004791:Enzyme Inhibitors; D008024:Ligands; D020543:Proteome; C113584:purvalanol B; D016203:CDC2 Protein Kinase; D018844:Cyclin-Dependent Kinases; D000225:Adenine