Single Molecule Spectroscopy of Amino Acids and Peptides by Recognition Tunneling

Abstract

The human proteome has millions of protein variants due to alternative RNA splicing and posttranslational modifications, and variants that are related to diseases are frequently present in minute concentrations. For DNA and RNA, low concentrations can be amplified using the polymerase chain reaction, but there is no such reaction for proteins. Therefore, the development of single molecule protein sequencing is a critical step in the search for protein biomarkers. Here we show that single amino acids can be identified by trapping the molecules between two electrodes that are coated with a layer of recognition molecules and measuring the electron tunneling current across the junction. A given molecule can bind in more than one way in the junction, and we therefore use a machine-learning algorithm to distinguish between the sets of electronic ‘fingerprints’ associated with each binding motif. With this recognition tunneling technique, we are able to identify D, L enantiomers, a methylated amino acid, isobaric isomers, and short peptides. The results suggest that direct electronic sequencing of single proteins could be possible by sequentially measuring the products of processive exopeptidase digestion, or by using a molecular motor to pull proteins through a tunnel junction integrated with a nanopore. The proteome is likely a much better molecular indicator of the current health status of humans than the genome,2 but proteomic data are harder to acquire.3 Protein sequences deduced from cDNA lack information about alternative splicing and post-translational Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use:http://www.nature.com/authors/editorial_policies/license.html#terms Correspondence and requests for material should be addressed to SL. *These authors contributed equally Author Contributions YZ and HL carried out tunneling measurements with assistance from SS, WS and JI. BA wrote the SVM code and analyzed data. BG contributed to the analysis. SM carried out force spectroscopy experiments. CB and SB carried out the electrospray MS. PZ and SL designed experiments and SL wrote the paper. Competing Financial Interests YZ, PZ and SL are named as inventors in patent applications. SL is cofounder of a company based on this technology. Additional Information Supplementary Information accompanies this paper. HHS Public Access Author manuscript Nat Nanotechnol. Author manuscript; available in PMC 2014 December 01. Published in final edited form as: Nat Nanotechnol. 2014 June ; 9(6): 466–473. doi:10.1038/nnano.2014.54. A uhor M anscript

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@inproceedings{Zhao2014SingleMS, title={Single Molecule Spectroscopy of Amino Acids and Peptides by Recognition Tunneling}, author={Yanan Zhao and Brian Alan Ashcroft and Peiming Zhang and Hao Liu and Suman K. Sen and Weisi Song and JongOne Im and Brett J Gyarfas and Saikat Kumar Manna and Sovan Biswas and Chad R Borges and Stuart Lindsay}, year={2014} }