Real-time dynamic single-molecule protein sequencing on an integrated semiconductor device
Brian D. Reed, Michael J. Meyer, Valentin Abramzon, Omer Ad, Omer Ad, Pat Adcock, Faisal R. Ahmad, Gün Alppay, James A. Ball, James H. Beach, Dominique Belhachemi, Anthony Bellofiore, Michael Bellos, Juan Felipe Beltrán, A.K. Betts, Mohammad Wadud Bhuiya, Kristin Blacklock, Robert E. Boer, D.C. Boisvert, Norman D. Brault, Aaron Buxbaum, Steve Caprio, Changhoon Choi, Thomas D. Christian, Robert Clancy, Joseph H. Clark, Thomas Connolly, Kathren Fink Croce, Richard Cullen, M. Smith Davey, Jack Davidson, Mohamed M. Elshenawy, Michael Ferrigno, Daniel Frier, Saketh Gudipati, Stephanie Hamill, Zhaoyu He, Sharath Hosali, Haidong Huang, Le Huang, Ali Kabiri, Gennadiy Kriger, Brittany Lathrop, An Li, Peter A.C. Lim, Stephen Liu, Feixiang Luo, Caixia Lv, Xiaoxiao Ma, Evan McCormack, Michele Millham, Roger R. Nani, Manjula Pandey, John Parillo, Gayatri Patel, Douglas H. Pike, Kyle Preston, Adeline Pichard-Kostuch, Kyle Rearick, Todd M. Rearick, Marco Ribezzi‐Crivellari, Gerard M. Schmid, Jonathan H. Schultz, Xinghua Shi, Badri Nath Singh, Nikita Srivastava, Shannon F. Stewman, T. R. Thurston, T. R. Thurston, Philip Trioli, Jennifer Tullman, Xin Wang, Yen‐Chih Wang, Eric A. G. Webster, Zhizhuo Zhang, Jorge Zuniga, Smita S. Patel, Andrew D. Griffiths, Antoine M. van Oijen, Michael McKenna, Matthew D. Dyer, Jonathan M. Rothberg
Abstract
Studies of the proteome would benefit greatly from methods to directly sequence and digitally quantify proteins and detect posttranslational modifications with single-molecule sensitivity. Here, we demonstrate single-molecule protein sequencing using a dynamic approach in which single peptides are probed in real time by a mixture of dye-labeled N-terminal amino acid recognizers and simultaneously cleaved by aminopeptidases. We annotate amino acids and identify the peptide sequence by measuring fluorescence intensity, lifetime, and binding kinetics on an integrated semiconductor chip. Our results demonstrate the kinetic principles that allow recognizers to identify multiple amino acids in an information-rich manner that enables discrimination of single amino acid substitutions and posttranslational modifications. With further development, we anticipate that this approach will offer a sensitive, scalable, and accessible platform for single-molecule proteomic studies and applications.