Final Report Summary - NANOFORBIO (Nanostructures for biology)
The NANOforBIO project employed our advanced capabilities for nanofabrication to explore new biology at the single-molecule and single-cell level. We specifically addressed two directions:
(i) Single biomolecule translocation through solid state nanopores
We developed and exploited solid-state nanopores for the study of real-time translocation of individual biomolecules. We explored screening of DNA, proteins, and DNA-protein complexes at the single-molecule level, and we will built biomimetic nanopores. We investigated the detection of local proteins bound to DNA with nanopores. After our initial work with RecA, we concentrated on a number of different protein systems, including nucleosomes. We first studied the translocation of free proteins. We succeeded in realizing biomimetic nanopores that mimic the nuclear pore complex. In particular we realized selectivity, where transport receptors proceeded through the pores, whereas the passage of non-specific proteins was strongly inhibited. Furthermore we developed a technique to insert a-haemolysin biological pores into solid-state nanopores. An exciting development is that we can use a single sheet of graphene, one single-atom thin, as a membrane for nanopores.
(ii) Nanostructures for bacteria
We use nanofabrication to create well-defined spatial confinements for bacteria for biophysical studies of the interaction between bacteria and their habitat with an unprecedented control of the spatial structure and habitat parameters. It allows to experimentally address a number of fundamental issues in the ecology and evolution of bacteria. We successfully used microfabrication of bacterial habitats to study ecology and evolution. We did, for example, use our nanostructures to study the mechanism to generate cooperation among bacteria and studied the effect of antibiotics. Additionally, microfabrication opened up a way to explore the biophysics of bacteria in confined space. We exploited the unique phenotype of bacteria in the nanofabricated slits to elucidate the mechanism of cell division. Specifically we studied Min oscillations in bacteria that were shape-sculpted by nanofabrication and initiated bottom-up in vitro studies of Min patterns in confined nanochambers.
The 5-year ERC grant was highly successful, as can be measured in various ways. It resulted in a number of exciting scientific breakthroughs (first graphene nanopores, first hybrid biological/solid-state nanopores, biomimetic nuclear pore complexes, discovery of DNA supercoil hopping, bacterial ecological games on chip, evidence for nucleoid occlusion in bacteria, discovery of DNA knots, first Min protein oscillations in shape-shifted bacteria). Furthermore, the results were published in 73 publications, including 35 in top journals (2 Science, 1 Cell, 8 Nature X, 24 in PNAS, Molecular Cell, Nano Letters, ACS Nano, etc).
(i) Single biomolecule translocation through solid state nanopores
We developed and exploited solid-state nanopores for the study of real-time translocation of individual biomolecules. We explored screening of DNA, proteins, and DNA-protein complexes at the single-molecule level, and we will built biomimetic nanopores. We investigated the detection of local proteins bound to DNA with nanopores. After our initial work with RecA, we concentrated on a number of different protein systems, including nucleosomes. We first studied the translocation of free proteins. We succeeded in realizing biomimetic nanopores that mimic the nuclear pore complex. In particular we realized selectivity, where transport receptors proceeded through the pores, whereas the passage of non-specific proteins was strongly inhibited. Furthermore we developed a technique to insert a-haemolysin biological pores into solid-state nanopores. An exciting development is that we can use a single sheet of graphene, one single-atom thin, as a membrane for nanopores.
(ii) Nanostructures for bacteria
We use nanofabrication to create well-defined spatial confinements for bacteria for biophysical studies of the interaction between bacteria and their habitat with an unprecedented control of the spatial structure and habitat parameters. It allows to experimentally address a number of fundamental issues in the ecology and evolution of bacteria. We successfully used microfabrication of bacterial habitats to study ecology and evolution. We did, for example, use our nanostructures to study the mechanism to generate cooperation among bacteria and studied the effect of antibiotics. Additionally, microfabrication opened up a way to explore the biophysics of bacteria in confined space. We exploited the unique phenotype of bacteria in the nanofabricated slits to elucidate the mechanism of cell division. Specifically we studied Min oscillations in bacteria that were shape-sculpted by nanofabrication and initiated bottom-up in vitro studies of Min patterns in confined nanochambers.
The 5-year ERC grant was highly successful, as can be measured in various ways. It resulted in a number of exciting scientific breakthroughs (first graphene nanopores, first hybrid biological/solid-state nanopores, biomimetic nuclear pore complexes, discovery of DNA supercoil hopping, bacterial ecological games on chip, evidence for nucleoid occlusion in bacteria, discovery of DNA knots, first Min protein oscillations in shape-shifted bacteria). Furthermore, the results were published in 73 publications, including 35 in top journals (2 Science, 1 Cell, 8 Nature X, 24 in PNAS, Molecular Cell, Nano Letters, ACS Nano, etc).