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Control and measurement of single macromolecules in space and time

Periodic Reporting for period 2 - COSMOS (Control and measurement of single macromolecules in space and time)

Reporting period: 2019-12-01 to 2021-05-31

We recently developed a new experimental approach to trapping a single molecule, such as a protein, in solution, based on its electrical charge. The proposal focuses on developing a novel measurement principle (escape time electrometry - ETe) for highly precise measurements of electrical charge on single molecules in the fluid phase. One of the main goals of the project is to use the ETe methodology to rapidly measure distributions in electrical charge of biomolecular species in solution. The potential application base is extremely broad with areas of immediate interest including detection of: (1) heterogeneous states of multimeric proteins, (2) antigen- Antibody interactions (3) small molecule binding to drug target proteins, (4) post-translational modifications such as phosphorylation, (5) conformational changes in biomolecules. The ETe measurement approach relies on optical observation of single molecules in a microscope and therefore requires the attachment of fluorescent labels to the molecule of interest. In a parallel sphere of activity we are also looking to develop the ability to develop an all electrical, label-free approach to the detection of the electrical properties of a single molecule in solution. The development of these new experimental approaches will not only enable researchers to address fundamental questions on the nature of molecular interactions and properties in solution but will also provide a new highly sensitive and precise biomedical detection technology that could have direct impact on aspects of societal health.
The implementation of the project in the first reporting period has been highly successful. My research group relocated from Zurich to Oxford as of 01.06.2018. The first few months of the project were devoted entirely to relocating and setting up the laboratory from scratch, re-building experimental set-ups and restarting experiments. One of the major fronts on which we had to invest significant effort was the transfer of our nanofabrication processes to the local cleanroom facilities. This enterprise is now nearing successful completion, but it has not come without fantastic team work and significant amounts of time and effort on the part of the group members funded by the ERC. The transition has been successful overall and our efforts are clearly beginning to bear fruit.

The recruited team started out with two doctoral (DPhil) students, and a postdoc working on theory and computation, all of whom transferred with the group from Zurich to Oxford. Within 6 to 9 months of moving to Oxford we had recruited two additional full-time postdocs and one part-time technician on the experimental side, and an additional PhD student with a focus on computation. The group currently consists of 6 doctoral students and 4 postdocs all contributing to various aspects of the project. The team now has the right strength and mix of skills, team members have acquired the relevant background training in our fields of interest and are poised to make a concerted move towards realising the goals of the project.

In the first phase of the project we have specifically been able to reproduce and improve upon previous electrometry measurements on single molecules in solution. In addition we already count a few major additional unpublished research achievements. We have: (1) demonstrated the ability to use electrometry to address questions related to molecular structure of nucleic acids like DNA and RNA, (2) developed a new experimental approach that facilitates the imaging of surface electrical charge distribution of material interfaces immersed in an electrolyte, and (3) made fundamental advances into the understanding key mechanisms underlying electrostatic interactions in solution. Overall, in the first phase of the project we have laid the foundations for breakthroughs on three fronts comprising new technologies, fundamentally new molecular measurement ability and basic science discovery.
By the end of the project we expect to have developed new technologies that will enable for the first time high precision measurement of molecular properties in the fluid phase. We anticipate the ETe approach will have developed into a mature experimental platform, and that we will have a proof-of-concept working prototype for an all-electrical measurement of the properties of a single molecule in the fluid phase. In delivering high precision measurements of molecular charge at high throughput and at the single molecule level, both the microscopy-based ETe approach and the all-electrical approach currently under development, constitute major progress in molecular measurement that are well beyond the state-of-the-art. We anticipate that the new opportunities for molecular measurement and detection that will emerge from the project will have significant impact both on fundamental research as well as on the biotechnology and pharmaceutical sectors. From our efforts on the COSMOS project, we further anticipate being able to count an unanticipated and far-reaching fundamental discovery on the nature of intermolecular interactions in solution.