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Department of Chemistry

Portrait of dk10012

Watching single molecules in action

We are physical scientists interested in developing and applying a range of new quantitative biophysical methods, based on single molecule  fluorescence  and scanning probe microscopy, to important problems in biology, which have not been addressed to date due to the lack of suitable tools and hence  directly image important and disease relevant  biological processes at  the level of single molecules. While our experiments range from experiments in  test-tubes to imaging  living cells, no previous biological background is required for our research since  we work closely with a range of biological and clinical collaborators. We are also part of the Cambridge Dementia Research centre.

Single molecule fluorescence

By studying molecules one at time, using fluorescence, specific complexes in a mixture can be identified and analysed without the need for any separation. With our collaborators we are exploiting single molecule fluorescence spectroscopy to study a range of biologically important molecules and processes. At present our main projects are:

  • Imaging the early molecular events that lead to the triggering of T-cells, a process that underpins the adaptive immune response and is remarkably  sensitive and selective.
  • Imaging the early molecular events that lead to the triggering of Toll-like receptors, a key process in the innate immune response which plays an important role in causing inflammation in neurodegenerative diseases such as Alzheimer’s disease.
  • Imaging and characterising the protein aggregates formed in the test-tube during aggregation reactions of disease-associated proteins and those present in human samples such as cerebral spinal fluid and brain tissue. We want to determine the aggregates responsible for causing neurodegenerative diseases such as Alzheimer’s and Parkinson’s disease in humans and the mechanisms by which they damage cells and spread through the brain.
  • Determining the structure and organisation of DNA in the nucleus of cells and  how this is regulated and changes during  cell differentiation.
  • Imaging aggregates of P53 and determining the role of these aggregates in the development and spreading of cancer.  
  • Developing and improving our imaging methods. At present our focus is developing methods to image protein aggregates at 20 nm spatial resolution and determine their composition and structure.

Scanning nanopipette

In collaboration with Professor Korchev at Imperial College we have developed a method based on a scanning nanopipette that allows robust, high resolution, non- contact imaging of living cells, down to the level of individual protein complexes. It can also be used to probe function by performing nanoscale assays such as locally deliver controlled amounts of reagents or performing single ion channel recording. The figure shows the University of Cambridge crest written in fluorescent DNA. We are using this method to watch the details of biological process taking place on the surface of living cells and to directly observe protein aggregates damage neuronal cells.


For a recent review  of our work on neurodegenerative disease see: Imaging individual protein aggregates to follow aggregation and determine the role of aggregates in neurodegenerative disease. Biochimica et Biophysica Acta - Proteins and Proteomics 1867,870 (2019).



Selected Publications

Different soluble aggregates of Abeta 42 can give rise to cellular toxicity through different mechanisms.Nat Commun 10,1541 (2019).

A cell-topography based mechanism for ligand discrimination by the T-cell receptor. PNAS 116,14002, (2019).

Activation of Toll-like receptors nucleates assembly of the MyDDosome signaling hubeLife 7, (2018) . 

Inhibiting the Ca2+ Influx Induced by Human CSFCell Rep 21, 3310 (2017)

Initiation of T cell signalling by CD45 segregation at "close contacts". Nature Immunology 17, 574 (2016).

Kinetic model of the aggregation of alpha-synuclein provides insights into prion-like spreading. PNAS 113, E1206 (2016).

A mechanistic model of tau amyloid aggregation based on the direct observation of oligomers. Nature Communication 6, 7025 (2015).

Local delivery of molecules from a nanopipette for quantitative receptor mapping on live cells. Analytical chemistry 85, 9333 (2013).

Ubiquitin chain conformation regulates recognition and activity of interacting proteins. Nature  492, 266-270 (2012).

Direct Observation of the Interconversion of Normal and Toxic Forms of alpha-Synuclein. Cell  149, 1048-1059 (2012).


MyD88 Death-Domain Oligomerization Determines Myddosome Architecture: Implications for Toll-like Receptor Signaling
MC Moncrieffe, D Bollschweiler, B Li, PA Penczek, L Hopkins, CE Bryant, D Klenerman, NJ Gay
– Structure
The Costs of Close Contacts: Visualizing the Energy Landscape of Cell Contacts at the Nanoscale.
K Kulenkampff, AH Lippert, J McColl, AM Santos, A Ponjavic, E Jenkins, J Humphrey, A Winkel, K Franze, SF Lee, SJ Davis, D Klenerman
– Biophysical Journal
N-Terminal Ubiquitination of Amyloidogenic Proteins Triggers Removal of Their Oligomers by the Proteasome Holoenzyme
Y Ye, D Klenerman, D Finley
– J Mol Biol
A General in Vitro Assay for Studying Enzymatic Activities of the Ubiquitin System
Y Zuo, BK Chong, K Jiang, D Finley, D Klenerman, Y Ye
– Biochemistry
Analysis of α-synuclein species enriched from cerebral cortex of humans with sporadic dementia with Lewy bodies.
JB Sanderson, S De, H Jiang, M Rovere, M Jin, L Zaccagnini, A Hays Watson, L De Boni, VN Lagomarsino, TL Young-Pearse, X Liu, TC Pochapsky, BT Hyman, DW Dickson, D Klenerman, DJ Selkoe, T Bartels
– Brain communications
Alpha synuclein aggregation drives ferroptosis: an interplay of iron, calcium and lipid peroxidation.
PR Angelova, ML Choi, AV Berezhnov, MH Horrocks, CD Hughes, S De, M Rodrigues, R Yapom, D Little, KS Dolt, T Kunath, MJ Devine, P Gissen, MS Shchepinov, S Sylantyev, EV Pavlov, D Klenerman, AY Abramov, S Gandhi
– Cell Death and Differentiation
High-resolution label-free 3D mapping of extracellular pH of single living cells.
Y Zhang, Y Takahashi, SP Hong, F Liu, J Bednarska, PS Goff, P Novak, A Shevchuk, S Gopal, I Barozzi, L Magnani, H Sakai, Y Suguru, T Fujii, A Erofeev, P Gorelkin, A Majouga, DJ Weiss, C Edwards, AP Ivanov, D Klenerman, EV Sviderskaya, JB Edel, Y Korchev
– Nat Commun
α-Synuclein strains target distinct brain regions and cell types.
A Lau, RWL So, HHC Lau, JC Sang, A Ruiz-Riquelme, SC Fleck, E Stuart, S Menon, NP Visanji, G Meisl, R Faidi, MM Marano, C Schmitt-Ulms, Z Wang, PE Fraser, A Tandon, BT Hyman, H Wille, M Ingelsson, D Klenerman, JC Watts
– Nat Neurosci
Different soluble aggregates of Aβ42 can give rise to cellular toxicity through different mechanisms
S De, DC Wirthensohn, P Flagmeier, C Hughes, FA Aprile, FS Ruggeri, DR Whiten, D Emin, Z Xia, JA Varela, P Sormanni, F Kundel, TPJ Knowles, CM Dobson, C Bryant, M Vendruscolo, D Klenerman
– Nature Communications
Soluble aggregates present in cerebrospinal fluid change in size and mechanism of toxicity during Alzheimer's disease progression
S De, DR Whiten, FS Ruggeri, C Hughes, M Rodrigues, DI Sideris, CG Taylor, FA Aprile, S Muyldermans, TPJ Knowles, M Vendruscolo, C Bryant, K Blennow, I Skoog, S Kern, H Zetterberg, D Klenerman
– Acta Neuropathologica Communications
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