Professor Mark D Szczelkun

The research focus of the group is the mechanistic analysis of bacterial defence systems that regulate Horizontal Gene Transfer, such as Restriction-Modification and CRISPR. In addition to their biologically relevant roles in influencing the acqusition of pathogenic and anti-microbial resistance genes, bacterial defences have also provided the basis of many important lab tools for manipulating DNA, such as the cloning and genome editing technologies.

We use a multidisciplinary experimental approach to studying DNA-protein interactions – combining single molecule microscopy (TIRF, magnetic tweezers, C-Trap), ensemble biochemistry (including millisecond time-resolution rapid-mixing fluorescence spectroscopy, molecular biology and protein chemistry) and Next Generation Sequencing (in particular, nanopore).

Chopping up DNA that invades a cell 

We have focussed our research efforts on Restriction-Modification enzymes that use ATP-dependent protein machines to cleave invading DNA into smaller fragments, addressing how these "molecular motors" convert chemical energy into mechanical events that lead to nuclease activity. We have been able to demonstrate alternative properties of the helicase-like motor domains of these enzymes, including dsDNA translocation and molecular switching. More recently we have started to explore the expanding range of defence systems that have been discovered from metagenomic analysis.  We aim to understand the diversity of these mechanisms, and their potential fitness costs to the bacteria. 

Chopping up DNA to allow gene editing

The Clustered, Regularly Interspaced, Short Palindromic Repeats (CRISPR) and the CRISPR-associated (cas) genes comprise an adaptive immune system in bacteria and archaea.  Silencing of foreign nucleic acids by CRISPR/Cas systems relies on a small CRISPR RNA (crRNA), the latter derived by processing transcribed CRISPR repeat-spacer arrays. We have developed a single molecule assay that allows the crRNA-guided recognition of specific DNA sequences to be followed in real time. Understanding how CRISPR/Cas systems achieve specificity will be particularly important in the manipulation of these proteins as tools for genome surgery, where specificity is paramount.