The nanomechanics of a single protein

Lead Research Organisation: King's College London
Department Name: Physics


Each organ in our body is composed of a large number of individual cells working together in a coordinated fashion. Inside each cell, there are thousands of different proteins that perform their function in a very well-established and synchronized way. In general, each of these proteins can be found in two different shapes -the folded and the unfolded states. Proteins unfold and refold continuously in our bodies once they are expressed in the ribosomes, which are the small factories where they are produced. Most proteins are 'active' or 'functional' only when they are in their folded state. Failing to fold gives rise to a myriad of devastating diseases such as Alzhemier's, Parkinson's, BSE (Mad Cow Disease) and many others. Therefore, we need experimental techniques able to track the folding routes of each individual protein undergoing a folding reaction to identify where and why each individual protein deviates from the 'correct' folding highway, being trapped at an intermediate state. This can be now be addressed by using state-of-the-art single molecule force-clamp spectroscopy. Using this approach, proteins are unfolded by the presence of a low (a few piconewtons) mechanical force, and once the force is reduced, the protein folds from highly extended states. Indeed, there are many proteins in our body that are continuously performing their function under the effect of a mechanical force. For example, the proteins involved in muscle elasticity, with crucial function also in e.g. the heart tissue, have to stretch and relax in a reversible way thousands of time every day. Failing to do that might have tragic consequences, resulting in muscle atrophy and, in the most severe cases, cardiac myopathies. Therefore, understanding how a mechanical force controls protein folding in these proteins is of capital importance, and it is far from being understood. In order to control muscle elasticity protein elasticity, nature has devised internal 'locks', called disulfide bonds, which prevent the protein to overstretch under high stress conditions. Such internal mechanical clamps can be mechanically 'open' through a covalent chemical reaction when required. Therefore, understanding the mechanisms to control these 'mechanical switches' is also of paramount importance in biophysics.
I will use the novel single molecule force-clamp spectroscopy technique to study the different trajectories followed by an unfolded protein in its journey to the native state. This technique has already proved successful at identifying, for the first time, the different conformations adopted by a protein that has been evolutionarily designed to fold within biological timescales. However, little is known about the mechanisms employed by 'mechanical proteins' to reversibly fold against a pulling force on a short timescale and without the intervention of energy spending mechanisms. I will investigate the conformational dynamics of a series of key proteins that control elasticity in the muscle, in the cytoskeleton and in the extracellular matrix. Next, I will study the effect of force on the reduction of a single disulfide bond embedded within the protein core. In particular, I will study how forces changes the outcome of a chemical reaction, and I will characterize the structure of the 'critical summit point' of the reaction, called transition state, which contains the relevant chemical information on the reaction outcome. Finally, I will examine how disulfide bonds affect the folding of a single protein, a phenomenon occurring in vivo to a wide variety of proteins composing the extracellular matrix. Altogether, these single molecule techniques have now reached a level of maturity where they can be used to attack more significant challenges in biology such as the basic biological mechanisms leading to protein protein and misfolding, especially in these proteins where preserving mechanical extensibility is key to maintain their physiological function.

Planned Impact

The gain in knowledge from the proposed research might have impact on the nation's health and wealth in several different ways. First, increase in basic knowledge about fundamental processes occurring in the human body, and understanding how and why diseases occur increased the cultural background of society. Secondly, controlling the outcome of a chemical reaction by applying mechanical force to the reagent system at the industrial scale might turn out to be a tour-de-force in some of the industries that deal with, e.g. particular polymer isomers. This could potentially have an important economic impact. On a deeper layer of knowledge, the increase in our basic knowledge increases the competitiveness of UK universities, thereby allowing them to attract more foreign students. Finally, staff working on the proposed research project will acquire research and professional skills that they might apply later in their careers, either as researchers, academics, or in the private industry field. It is expected that the PDRAs will present their work at numerous international conferences and in results in high impact scientific journals. King's College London is also committed to ensuring that research staff and students develop research, vocational and entrepreneurial skills that are matched to the demands of their future career paths. Both the Department of Physics and the Randall Division are a multidisciplinary environment and as such also provides an excellent place for training researchers at any level in new techniques.
We plan to engage the beneficiaries in several different ways. First, we will present the on-going research in three already scheduled public lectures. The first will take place in the framework of the London Structural Biology meetings organized by UCL. The second will take place in a soiree of the London Thomas Young Centre for Theory and Simulation of Materials, and the third in the specialized and world-reputed Gordon Conference on Single Molecules. Second, we plan to organize a workshop in single molecule mechanics. The PI (Dr. Garcia-Manyes) co-organized in 2010 a workshop in Bilbao, Spain, where students from the whole Europe had the opportunity to perform hands-on experiments and to analyse their own data during an intensive week of work. The plan is now to repeat this workshop in central London. Finally, the Public Relations Department at King's College London regularly publishes press releases reporting the results of its researchers; this information is communicated through the Web and through magazines, newsletters and annual reports for the general public, which are widely distributed. All these media channels will be used as appropriate and feasible to communicate the results of the present project to the public. School pupils attend our Open Days and undertake work experience in our laboratories.
Beneficiaries in the biotechnology and materials science industry will be engaged through King's College London Business Ltd. That provides a getaway to access the wealth of knowledge and expertise available at King's College London. KCL Business has extensive experience to create significant partnerships with industry to maximize innovation and to add value to our partner's business goals and help increase their global effectiveness. KCL Business offers access to knowledge and people, consultancy, collaborative and contract research, the opportunity to develop and licence new intellectual property and to engage with newly created spin-out companies through partnership and investment. If new discoveries will be made during the course of the proposed research, the PI will liaise with KCL Business to assess potential exploitability of the discovery before any publication. KCL Business is well experienced in providing advice in intellectual property rights and will ensure successful protection of intellectual property by patenting novel inventions and discoveries.
Description We have recently measured, for the first time, the mechanochemical properties of a copper-sulfur bond and a zinc/sulfur bond- We discovered two alternative chemical pathways to trigger non-enzymatic oxidative folding; me uncovered the mechanisms of mechanotrasduction of transcription facotrs through the nuclear pore complex; we developed a system to study the dynamics of breakthorugh of stacked lipid bilayers; one bilayer at a time
Exploitation Route In the general field of mechanochemistry
Sectors Chemicals,Healthcare,Manufacturing, including Industrial Biotechology

Description We have been invited to two high-end review articles, one in Nature Reviews Chemistry ( and another one in Nature Reviews Materials (under review) Our work has created a clear 'interdisciplinary impact', working across disciplines, and in particular focusing on the impact of physical sciences on biology. The development of single molecule and single cell measuring techniques - has had a direct impact on the biological sciences - and in particular for the molecular and cell biology fields. We have also started a collaboration with the instrument company Lumicks, resulting in an ICASE award
First Year Of Impact 2020
Sector Manufacturing, including Industrial Biotechology
Impact Types Societal

Description European Commission (Marie Curie program, IEF)
Amount £181,717 (GBP)
Funding ID 329308 
Organisation Marie Sklodowska-Curie Actions 
Sector Charity/Non Profit
Country Global
Start 03/2014 
End 03/2016
Description H2020-FETPROACT-2016-2017 (FETPROACT-2016)
Amount € 7,134,929 (EUR)
Funding ID 731957 
Organisation European Commission 
Department Horizon 2020
Sector Public
Country European Union (EU)
Start 01/2017 
End 01/2022
Description LINC-ing nanomechanics to gene expression: a single molecule approach
Amount £199,000 (GBP)
Funding ID RPG-2015-225 
Organisation The Leverhulme Trust 
Sector Charity/Non Profit
Country United Kingdom
Start 03/2016 
End 02/2019
Description Mechanochemical biology: a force to be reckoned with
Amount £942,614 (GBP)
Organisation The Leverhulme Trust 
Sector Charity/Non Profit
Country United Kingdom
Start 12/2017 
End 11/2022
Description Single Molecule insights into Nuclear Mechanotransduction
Amount £1,337,758 (GBP)
Funding ID 212218/Z/18/Z 
Organisation Wellcome Trust 
Sector Charity/Non Profit
Country United Kingdom
Start 01/2019 
End 12/2025
Description unveiling the molecular mechanisms underlying the onset of gamma-D-crystallin aggregation studied by single molecule force-clamp spectroscopy
Amount £99,290 (GBP)
Funding ID 1562 
Organisation Fight for Sight 
Sector Charity/Non Profit
Country United Kingdom
Start 09/2015 
End 09/2018