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Molecular machines based on coiled-coil protein origami

Periodic Reporting for period 2 - MaCChines (Molecular machines based on coiled-coil protein origami)

Reporting period: 2020-03-01 to 2021-08-31

Proteins are most versatile biopolymers that underlay functions and structures of all living organisms. Natural proteins form a wide range of different structures with a large variety of functions; however natural proteins represent only a tiny fraction of all possible polypeptide structures. The challenge is how to design new proteins that have not evolved in nature based on alternative design principles that may have new interesting features. Within the project MaCChines we aim to investigate and explore the potentials of designed proteins based on coiled-coil building modules (Coiled-coil protein origami CCPO) and take advantage of the unique properties of this platform.
Those programmable molecules have many properties that could be important for our society – they can be produced by cell factories in an energy and resource effective and sustainable manner, due to their structure defined at the nanoscale we can expect them to be efficient in recognition, delivery, catalysis and to find applications in medicine, biotechnology and other fields. It is likely that we are moving into an era of living and selfassembling machines that will find their use in all branches of technology.
The objective of MaCChines is to build foundations of a new branch of protein design science, develop and exploit unique features of de novo designed coiled-coil protein origami (CCPO), in order to enable new drug delivery technologies, construction of complex nanoscaffolds, new materials, sensors, rational design of molecular machines and new advanced therapeutics.
Within the first half of the project we can report advances on several exciting directions along the proposed directions of the ERC AdG MaCChines project and also along some new ideas that have been conceived and realized during this project.
We have developed several types of functionalized CC modules and implemented them for different functions:
- New metal and pH dependent coiled—coil modules (Aupič et al., 2018)
- Fusion of CC modules with split proteases to generate fast responsive cellular logic (Fink et al., NatChemBiol 2019)
- Application of CC modules for modulation of localization and regulation of biochemical/biological processes (Lebar et al., 2020)
We introduced several new strategies of CCPO assembly:
- Multichain CCPO assembly (Lapenta et al., 2021)
- Design of the designed protein folding pathway and use of multiple copies of the same type of the CC peptide (Aupič et al., 2021)
Results generated within MaCChines so far introduced several innovative concepts in protein design that could be applied also to the design of other programmable polymers. Demonstration and concepts for the introduction of coiled-coil based modules into mammalian cells represent a foundation for their wider use by other researchers and for diverse applications. In the second half of the project I expect to introduce several new types of building modules that offer new possibilities of the design, both for the in vitro as well as for implementation within cells that could offer an unprecedented ability to control function of natural components of cells.
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