Surface plasmon resonance imaging-based protein arrays for high-throughput screening of protein-protein interaction inhibitors.
The E7 protein produced by high-risk human papillomavirus (HPV) induces a degradation of the retinoblastoma tumor suppressor RB through direct interaction, which suggests that an inhibitor for the interaction can be a potential anticancer drug. A surface plasmon resonance (SPR) imaging-based protein array chip was developed for the high-throughput screening of inhibitor molecules targeting RB-E7 interaction. The glutathione S-transferase-fused E7 protein (GST-E7) was first layered onto a glutathionylated gold chip surface that had been designed to specifically bind to GST-fused proteins. Subsequently, a microarrayer was used to spot the hexa-histidine-tagged RB proteins (His(6)-RB) onto the GST-E7-layered gold chip surface, and the resulting SPR image was analyzed. Upon increased His(6)-RB concentration in the spotting solution, the SPR signal intensity increased proportionally, indicating that His(6)-RB bound to GST-E7 in a concentration-dependent manner. The His(6)-RB/GST-E7 interaction was challenged by spotting the His(6)-RB solution in the presence of a RB binding peptide (PepC) derived from a motif on E7. The SPR imaging data showed that PepC inhibited the His(6)-RB/GST-E7 interaction in a concentration-dependent manner. Our results show that the SPR imaging-based protein array chip can be applied to screen small molecule inhibitors that target protein-protein interaction.
Jung
Sun Ok
SO
BioNanotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology, Yuseong, Daejeon.
Ro
Hyeon-Su
HS
Kho
Byung Hoon
BH
Shin
Yong-Beom
YB
Kim
Min Gon
MG
Chung
Bong Hyun
BH
D050725:Papillomavirus E7 Proteins / Q000737:chemistry*; D010446:Peptide Fragments / Q000737:chemistry; D040081:Protein Array Analysis / Q000379:methods*; D011506:Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry*; D016160:Retinoblastoma Protein / Q000737:chemistry*; D020349:Surface Plasmon Resonance / Q000379:methods*
D050725:Papillomavirus E7 Proteins; D010446:Peptide Fragments; D011506:Proteins; D016160:Retinoblastoma Protein