Surface plasmon resonance imaging protein arrays for analysis of triple protein interactions of HPV, E6, E6AP, and p53.
We have developed a surface plasmon resonance (SPR)-based protein microarray to study protein-protein interactions in a high-throughput mode. As a model system, triple protein interactions have been explored with human papillomaviral E6 protein, tumor suppressor p53, and ubiquitin ligase E6AP. Human papillomavirus (HPV) is known to be a causative agent of cervical cancer. Upon infection, the viral E6 protein forms a heterotrimeric protein complex with p53 and E6AP. The formation of the complex eventually results in the degradation of p53. In the present study, a GST-fused E6AP protein was layered onto a glutathione (GSH)-modified gold chip surface. The specific binding of GST-E6AP protein onto the gold chip surface was facilitated through the affinity of GST to its specific ligand GSH. The interacting proteins (E6 and/or p53) were then spotted. Detection of the interaction was performed using a SPR imaging (SPRI) technique. The resulting SPRI intensity data showed that the protein-protein interactions of E6AP, E6, and p53 were detected in a concentration-dependent manner, suggesting that the SPRI-based microarray system can be an effective tool to study protein-protein interactions where multiple proteins are involved.
Ro
Hyeon-Su
HS
Department of Microbiology and Institute of Life Science, Gyeongsang National University, Gajwa-Dong, Gyeongnam, Korea.
Koh
Byung Hoon
BH
Jung
Sun Ok
SO
Park
Hyun Kyu
HK
Shin
Yong-Beom
YB
Kim
Min-Gon
MG
Chung
Bong Hyun
BH
D006801:Humans; D018625:Microscopy, Atomic Force; D009856:Oncogene Proteins, Viral / Q000378:metabolism*; D027383:Papillomaviridae / Q000378:metabolism*; D040081:Protein Array Analysis / Q000379:methods; D025941:Protein Interaction Mapping / Q000379:methods; D020349:Surface Plasmon Resonance / Q000379:methods; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism*
C466080:E6 protein, human papillomavirus type 1; D009856:Oncogene Proteins, Viral; D016159:Tumor Suppressor Protein p53; C471639:UBE3A protein, human; D044767:Ubiquitin-Protein Ligases