DNA is the molecule that nature uses as genetic material and it rarely exists in a relaxed state inside cells. Rather, DNA is subjected to mechanical deformations like torsion, bend or stretch, which are induced by proteins and which facilitate the realization of all the different genetic transactions such as recombination, gene expression and replication. In particular, the formation of closed DNA loops is directly associated with the regulation of genes.
Nucleoid-associated proteins (NAPs) are a collection of DNA-interacting proteins that perform crucial roles of organization, packaging, and gene regulation in prokaryotic chromosomes. They often bind non-specifically and create a wide variety of DNA topologies. Integration host factor (IHF) is a key NAP in Escherichia coli and other Gram-negative bacteria. Its architectural role is thought to involve creating some of the sharpest bends observed in DNA, in excess of 160 ̊, thereby facilitating the assembly of higher-order nucleoprotein complexes such as gene regulatory loops, the CRISPR-Cas system, the origin of replication, and the Holliday junction. IHF’s large repertoire of roles supports the long-standing view that it has an essential function in the structural organization of DNA in a wide variety of genetic processes.
The relevance of DNA not only comes from biology but also lately has been applied in the field of nanotechnology. The remarkable specificity of the interactions between complementary bases makes DNA a construction material that can be used in the design of nano-architectures such as drug carriers or artificial molecular machines such as DNA tweezers or DNA walkers. However, DNA has other degrees of freedom, apart from the ones related with the specific base-pairing rules, that have not been exploited for engineering, including, for example, torsion and bending.
The aim of this project is to exploit the DNA bending induced by IHF for designing unique and exclusive DNA topologies. We plan to study which is the DNA bending angle that IHF imposes depending on different binding sites. We then plan to choose the one that causes a more bistable mechanical switch on the DNA so we can program the formation of closed DNA loops.
In our previous study, we compared atomic force microscopy (AFM) with all-atom molecular dynamics simulations (MDS) for the same DNA constructs of around 300 bp (1). New developments on simulation and microscopy imaging developed at the respective labs of Agnes Noy and Mark Leake allow the combination of both techniques in the nano-scale, relevant on the DNA technology, for identifying the complex topological states that encompass the interactions between IHF and DNA (1).
The current project is an extension of this previous manuscript (1) and it will involve the following series of tasks:
1) The simulation of the IHF-DNA complex in different binding sites using the recently established protocol adapted for standard modelling packages like AMBER
2) The learning and application of advanced sampling modelling techniques for finding out the underlying free energy landscape on the different situations
3) The possibility to undertake AFM imaging and analysis for testing the predictions made by modelling
The success of this project would enable us to design DNA constructs for testing different biological hypothesis related with the role of DNA architecture in genomic function and would give a total new dimensionality to DNA technology
1. S Yoshua, G Watson, J Howard, V Velasco-Berrelleza, MC Leake, A Noy (2020). “A nucleoid-associated protein bends and bridges DNA in a multiplicity of topological states with varying specificity” Nuc Acids Res, 49,8684-8698
How to apply:
Applicants should apply via the University’s online application system at https://www.york.ac.uk/study/postgraduate-research/apply/. Please read the application guidance first so that you understand the various steps in the application process.
Funding Notes
This is a self-funded project and you will need to have sufficient funds in place (eg from scholarships, personal funds and/or other sources) to cover the tuition fees and living expenses for the duration of the research degree programme. Please check the School of Physics, Engineering and Technology website https://www.york.ac.uk/physics-engineering-technology/study/funding/ for details about funding opportunities at York.