Antibacterial nanopores composed of DNA
Lead Research Organisation:
University College London
Department Name: Chemistry
Abstract
Strategic Research Priority: Industrial biotechnology and bioenergy
Due to the alarming rise in antibiotic resistance and decline of active compounds, the development of new powerful bacteria-killing agents is of great importance. In this project, we will create and test bilayer-puncturing nanopores that lyse bacteria but leave eukaryotic cells unharmed. The nanopore will be composed of folded DNA and carry lipid anchors for membrane insertion. The synthetic nanopores will be characterized and examined to unterstand their selective interaction with bacterial but not human cell membranes. Synthetic DNA nanopores that mimic biological behaviour and insert into membranes have been prepared previously and attracted considerable scientific interest. But the use of synthetic pores to selectively kill bacteria is a completely new observation.
The project falls with the BBSRC's strategic priorities and covers topics of synthetic biology, nanobiotechnology, chemcial biology, microbiology, and biophysics. By developing novel antibacterial compounds.
The project is experimentally feasible in the time frame as it is supported by strong preliminary data generated jointly by the PI's group at UCL and the non-academic partner MR at NPL. The data show that DNA pores of 7 nm height and 5 nm width are active against both Gram-positive and Gram-negative pathogenic bacteria. Remarkably, activity is selective against bacteria but not human cells.
While the findings are scientifically striking and of clear biological impact, there are several fundamental questions that have to be answered within the project in order to clarify the science and help exploit it:
(i) What are the structural and chemical components of the pore that are essential for cell killing? The data suggest that the structure of a pore AND lipid anchors are essential. But can a minimal pore be constructed with the same killing activity? What is the influence of the number and position of lipid anchors on activity?
(ii) What is the mechanism of bacterial cell killing? Do the pores puncture an otherwise intact membrane or completely rupture it? What are the kinetics of the membrane binding and bacterial killing?
(iii) What is the reason for the selectivity of the pores? Do they recognise membrane components of the bacterial membrane which are not present in eucaryotic cells?
(iv) Can bacteria-killing DNA pores be developed further and serve as conceptual template to create related nanomaterials or organic small-molecule drugs that achieve selective killing?
The proposed project is relevant to the non-academic partner, who has a long-standing and strategic interest in developing advanced measurement approaches, materials and methods to address the problem of antimicrobial resistance.
The elibility of the non-academic partner NPL has been confirmed with Dr. Nadine Mogford.
The projects is supported by a strong track-record of the SH and MR in the relevant areas (see point 4 below) and benefits from previous joint collaboration between SH and MR in the supervision of a joint PhD student.
Due to the alarming rise in antibiotic resistance and decline of active compounds, the development of new powerful bacteria-killing agents is of great importance. In this project, we will create and test bilayer-puncturing nanopores that lyse bacteria but leave eukaryotic cells unharmed. The nanopore will be composed of folded DNA and carry lipid anchors for membrane insertion. The synthetic nanopores will be characterized and examined to unterstand their selective interaction with bacterial but not human cell membranes. Synthetic DNA nanopores that mimic biological behaviour and insert into membranes have been prepared previously and attracted considerable scientific interest. But the use of synthetic pores to selectively kill bacteria is a completely new observation.
The project falls with the BBSRC's strategic priorities and covers topics of synthetic biology, nanobiotechnology, chemcial biology, microbiology, and biophysics. By developing novel antibacterial compounds.
The project is experimentally feasible in the time frame as it is supported by strong preliminary data generated jointly by the PI's group at UCL and the non-academic partner MR at NPL. The data show that DNA pores of 7 nm height and 5 nm width are active against both Gram-positive and Gram-negative pathogenic bacteria. Remarkably, activity is selective against bacteria but not human cells.
While the findings are scientifically striking and of clear biological impact, there are several fundamental questions that have to be answered within the project in order to clarify the science and help exploit it:
(i) What are the structural and chemical components of the pore that are essential for cell killing? The data suggest that the structure of a pore AND lipid anchors are essential. But can a minimal pore be constructed with the same killing activity? What is the influence of the number and position of lipid anchors on activity?
(ii) What is the mechanism of bacterial cell killing? Do the pores puncture an otherwise intact membrane or completely rupture it? What are the kinetics of the membrane binding and bacterial killing?
(iii) What is the reason for the selectivity of the pores? Do they recognise membrane components of the bacterial membrane which are not present in eucaryotic cells?
(iv) Can bacteria-killing DNA pores be developed further and serve as conceptual template to create related nanomaterials or organic small-molecule drugs that achieve selective killing?
The proposed project is relevant to the non-academic partner, who has a long-standing and strategic interest in developing advanced measurement approaches, materials and methods to address the problem of antimicrobial resistance.
The elibility of the non-academic partner NPL has been confirmed with Dr. Nadine Mogford.
The projects is supported by a strong track-record of the SH and MR in the relevant areas (see point 4 below) and benefits from previous joint collaboration between SH and MR in the supervision of a joint PhD student.
Studentship Projects
| Project Reference | Relationship | Related To | Start | End | Student Name |
|---|---|---|---|---|---|
| BB/M009513/1 | 01/10/2015 | 31/03/2024 | |||
| 1764872 | Studentship | BB/M009513/1 | 01/10/2016 | 30/03/2021 | Conor Lanphere |
| Description | A DNA nanopore has been designed that has a lid, which features an aptmer. This aptmer is selective for a specific protein, which once bound, opens the lid and activates the pore. The aptmer chosen here is a previously published human-alpha thrombin binding aptmer; however, this is the first time it has been incorporated onto a DNA nanopore. The controlled DNA nanopore was also used as part of a nano devise that was able to controllably release a drug and reduced cell viability 150% more than the drug on its own in addition to remove off target effects. A DNA nanopore composed to two pieces has been designed that can undergo triggered fusion to act as a potential synthetic ion channel. This further allows DNA nanopores to be controlled in their application and further reduce off-target effects. Enrichment on a membrane surface was investigated looking at how steric affect DNA hybridisation. This work will allow us to enrich nanopores on cells surfaces in an effort to increase their potency. |
| Exploitation Route | The two controlled DNA nanopores have potential as drug delivery vehicles. This has already been demonstrated in the case of the aptmer-gated pore, but only for one drug and one target. This could be tested for a number of different therapies. In addition, the aptmer can be readily exchanged to a different sequence such that the trigger can be changed without much difficulty allowing its application to be tailored. The synthetic ion channel has the potential to be a therapeutic in its own right and could be used to target diseases where ion channels become defunct. In addition, the triggered nature of the pore could be expanded to pores of different sizes or instead of a simple lock and key mechanism using a DNA-toehold this could be functionalized with an aptmer to form a pore only in the presence of a target. The effort to pre-enrich DNA nanopores on cells membranes could have wide ranging effects from improving the previously demonstrated cytotoxic response of DNA nanopores to improving their activity as antibacterial agents (currently this has not been demonstrated). This could also be used more generally to build DNA nanostructure on the surfaces of membranes for a wide range of potential applications. |
| Sectors | Healthcare,Pharmaceuticals and Medical Biotechnology |
| Description | Developed a set of protocols for the Design, Assembly and Characterization of DNA nanopores for Nature protocols |
| Geographic Reach | Multiple continents/international |
| Policy Influence Type | Influenced training of practitioners or researchers |
| Description | Collaboration with NPL as part of an iCASE PhD Studentship. |
| Organisation | National Physical Laboratory |
| Country | United Kingdom |
| Sector | Academic/University |
| PI Contribution | Use of the NPL facilities and expertise to advance science as part of the NPL remit. Concrete advances have not yet been achieved. |
| Collaborator Contribution | Use of the NPL facilities and expertise to advance the research of this project. |
| Impact | No outputs or outcomes have yet been achieved that are worth commenting on. |
| Start Year | 2016 |
| Description | Poster Presentation at RSC Nucleic Acids Forum |
| Form Of Engagement Activity | A talk or presentation |
| Part Of Official Scheme? | No |
| Geographic Reach | National |
| Primary Audience | Professional Practitioners |
| Results and Impact | Attended, along with several members of my group, the Nucleic Acids Forum at The Royal Society of Chemistry at Burlington House. I presented a poster on my work as part of the poster session. |
| Year(s) Of Engagement Activity | 2019 |