Daven Henze discusses how air pollution spreads across the globe and what policymakers are doing in response.
New Research In
Physical Sciences
Featured Portals
Articles by Topic
Biological Sciences
Featured Portals
Articles by Topic
Edited by Alan R. Fersht, Gonville and Caius College, Cambridge, United Kingdom, and approved February 21, 2018 (received for review May 1, 2017)
Significance
Protein sequencing remains a challenge for small samples. A sensitive sequencing technology will create the opportunity for single-cell proteomics and real-time screening for on-site medical diagnostics. To resolve protein identity, we previously developed a computational algorithm that analyzes the ordered sequence of only two types of amino acids within a protein species. Through modification of a biological nanomachine, here, we developed single-molecule fluorescence technology to linearize protein molecules and to read signals from labeled amino acids in an ordered manner. This proof of concept of single-molecule fingerprinting will open the door to single-molecule protein sequencing and pave the road toward the development of a new, fast, and reliable diagnostic tool.
Abstract
Proteomic analyses provide essential information on molecular pathways of cellular systems and the state of a living organism. Mass spectrometry is currently the first choice for proteomic analysis. However, the requirement for a large amount of sample renders a small-scale proteomics study challenging. Here, we demonstrate a proof of concept of single-molecule FRET-based protein fingerprinting. We harnessed the AAA+ protease ClpXP to scan peptides. By using donor fluorophore-labeled ClpP, we sequentially read out FRET signals from acceptor-labeled amino acids of peptides. The repurposed ClpXP exhibits unidirectional processing with high processivity and has the potential to detect low-abundance proteins. Our technique is a promising approach for sequencing protein substrates using a small amount of sample.
Footnotes
- ↵1To whom correspondence may be addressed. Email: anne{at}annemeyerlab.org or c.joo{at}tudelft.nl.
Author contributions: J.v.G., A.S.M., and C.J. designed research; J.v.G., M.F., M.S., and P.T. performed research; J.v.G. and M.F. analyzed data; and J.v.G., M.S., A.S.M., and C.J. wrote the paper.
Conflict of interest statement: J.v.G., C.J., and A.S.M. hold a patent (“Single molecule protein sequencing”; WO2014014347).
This article is a PNAS Direct Submission.
This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.1073/pnas.1707207115/-/DCSupplemental.
Published under the PNAS license.
Single-molecule peptide fingerprinting
Sign up for Article Alerts
Jump to section
You May Also be Interested in
Some researchers are aiming to apply the technique to a range of hard-to-treat maladies, including severe obsessive compulsive disorder, Tourette syndrome, and Alzheimer’s disease.
Image credit: Helen Mayberg.
Can a computer actually produce something new rather than merely emulating human artists? If nothing else, they are making some types of art more accessible for would-be artists.
Image credit: Roman Verostko (artist).
PNAS Profile of NAS member and geologist Paul E. Olsen.
Image courtesy of Kevin Krajick (Columbia University, New York).
Fossil evidence helps address a longstanding debate on the evolution of hagfish, a jawless, marine-dwelling slime “eel,” and suggests that living jawless vertebrates may not be as primitive as their anatomy suggests.
Image courtesy of Tetsuto Miyashita.
More Articles of This Classification
Biological Sciences
Related Content
Cited by...
- No citing articles found.