Cantilever sensor for nanomechanical detection of specific protein conformations.
A fast, label-free, and multiplexed method based on piezoresistive cantilevers is reported for the detection of specific protein conformations at the nanoscale level. The ligand-binding domain of the human oestrogen receptor (ERalpha-LBD) is used as the experimental model system, and ERalpha-LBD with or without oestradiol (E2) is detected using the conformation-specific peptides alpha/betaI (Ser-Ser-Asn-His-Gln-Ser-Ser-Arg-Leu-Ile-Glu-Leu-Leu-Ser-Arg, which recognizes E2-bound ER) and alpha/betaII (Ser-Ala-Pro-Arg-Ala-Thr-Ile-Ser-His-Tyr-Leu-Met-Gly-Gly, which recognizes E2-free ER). Target-specific signals are obtained in situ at protein concentrations of 2.5-20 nM. The in-build electrical readout of the piezoresistive cantilevers provides a convenient alternative to the conventional optical detection, and the presented method offers the possibility of detecting protein conformational changes using miniaturized microarrays.
Mukhopadhyay
Rupa
R
Interdisciplinary Nanoscience Center, Department of Physics and Astronomy, University of Aarhus, 8000 Aarhus C, Denmark. rupa@inano.dk
Sumbayev
Vadim V
VV
Lorentzen
Martin
M
Kjems
Jørgen
J
Andreasen
Peter A
PA
Besenbacher
Flemming
F
D015374:Biosensing Techniques / Q000295:instrumentation* / Q000379:methods; D004563:Electrochemistry / Q000295:instrumentation* / Q000379:methods; D004867:Equipment Design; D019544:Equipment Failure Analysis; D004958:Estradiol / Q000737:chemistry*; D047628:Estrogen Receptor alpha / Q000032:analysis / Q000737:chemistry*; D019563:Mechanics; D008832:Microchemistry / Q000295:instrumentation / Q000379:methods; D036103:Nanotechnology / Q000295:instrumentation* / Q000379:methods; D040081:Protein Array Analysis / Q000295:instrumentation* / Q000379:methods; D011485:Protein Binding; D011487:Protein Conformation; D025941:Protein Interaction Mapping / Q000295:instrumentation* / Q000379:methods; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D047628:Estrogen Receptor alpha; D004958:Estradiol