Detection of Bax protein conformational change using a surface plasmon resonance imaging-based antibody chip.
We describe an antibody chip technology that uses a surface plasmon resonance (SPR) imaging system to examine the conformational change of a protein. In this study, we used Bax protein, a pro-apoptotic member of the Bcl-2 family of proteins, as a model protein to investigate the conformational alteration triggered by a TNF-related apoptosis-inducing ligand (TRAIL), a potent inducer of apoptosis. To develop the antibody chip for detecting the Bax conformational change, we immobilized Bax monoclonal antibody 6A7, which recognizes only a conformationally changed Bax protein on a gold surface. The resultant immobilized Bax antibodies provided specific and accurate measurements of the active conformation-specific epitope in the apoptotic cancer cells treated with the TRAIL; these measurements corresponded to the data obtained by immunoprecipitation analysis using an active conformation-specific Bax antibody (6A7). The results of our study indicated that TRAIL-induced Bax structural change could be monitored quickly and simply using an SPR imaging system, thus demonstrating the potential for using such a system for the analysis of conformational properties of target proteins.
Kim
Moonil
M
BioNanotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology, P.O. Box 115, Yuseong, Daejeon 305-600, Republic of Korea.
Jung
Sun Ok
SO
Park
Kyoungsook
K
Jeong
Eun-Ju
EJ
Joung
Hyou-Arm
HA
Kim
Tae-Hyoung
TH
Seol
Dai-Wu
DW
Chung
Bong Hyun
BH
D000906:Antibodies; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology; D051017:Apoptosis Regulatory Proteins / Q000494:pharmacology; D045304:Cytochromes c / Q000378:metabolism; D006801:Humans; D008562:Membrane Glycoproteins / Q000494:pharmacology; D040081:Protein Array Analysis; D011487:Protein Conformation / Q000187:drug effects; D021381:Protein Transport; D020349:Surface Plasmon Resonance / Q000379:methods*; D053221:TNF-Related Apoptosis-Inducing Ligand; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D051028:bcl-2-Associated X Protein / Q000737:chemistry* / Q000276:immunology
D000906:Antibodies; D051017:Apoptosis Regulatory Proteins; D008562:Membrane Glycoproteins; D053221:TNF-Related Apoptosis-Inducing Ligand; C505653:TNFSF10 protein, human; D014409:Tumor Necrosis Factor-alpha; D051028:bcl-2-Associated X Protein; D045304:Cytochromes c