Principal Investigator: Dr. Carlos Penedo

 

Single-molecule Biophysics

We are continuously trying to improve and develop new methods for single molecule fluorescence detection. Our efforts are currently focused in the following main areas: 1. New dyes with improved photophysics, brighter, more photostable and with less cross-talk and more defined spectral and temporal properties. 2. We are also interested in new and simpler methods for reversible surface immobilization 3. New fluorescent labeling strategies and ortoghonal chemistries for single-molecule FRET studies where a fluorescent donor and acceptor species can be unambiguously placed at specific positions within the biomolecular structure

 

Fighting bacterial pathogens: Functional dynamics of RNA motifs involved in gene regulation

Riboswitches are non-coding mRNA motifs found mostly in bacteria and fungi that control gene expression by sensing the concentration of a particular metabolite. Since their discovery in 2002, more than 15 riboswitch families sensing nucleotides, aminoacids, coenzymes, fluoride ions and many others have been discovered. To achieve their characteristic high specificity and selectivity towards their respective metabolites, riboswitches need to adopt a very defined 3D structure in a process known as folding. We apply single molecule techniques to understand this folding process. Our long-term aims are to use this information to develop metabolite analogs as antibacterial drug targets and also from a synthetic biology perspective to engineer riboswitches working as logic gates than can artificially regulate bacterial gene expression. Currently we are working on the adenine, SAM, lysine and fluoride riboswitches, although new ones such as the THF riboswitch are continuously being added to the list. In this project we closely collaborate with Prof. Daniel A. Lafontaine at the University of Sherbrooke (Quebec, Canada).

 

Protein misfolding and aggregation processes involved in neurological disorders

The b-amyloid peptide is formed inside the human brain from the proteolytic cleavage of the amyloid precursor protein. Naturally-occurring aggregated structures of these peptides can adopt a variety of coexisting morphologies ranging from small oligomers to the fibrils and plaques characteristic of Alzheimer’s disease. Unveiling their structural polymorphism is of crucial importance to fight against the growing impact of dementia-related disorders. We are developing fluorescence sensing methods capable ofreporting the major aggregated species present in solution. In combination with single-molecule studies we try to understand the mechanistic details of how these peptides unfold and aggregate and how this process is modulated by the surrounding environment. Another areas of interest include testing for small-molecule amyloid inhibitors and more recently the effect small heat shock proteins ont the aggregation process.

 

Molecular machines involved in RNA and DNA processing pathways

We are interested in understanding how complex molecular machines process DNA and RNA structures. Vital cellular tasks such as replication, transcription, RNA splicing and degradation rely on the interaction between multi-subunit protein complexes in which each protein has a defined function and a set of interacting partners. In collaboration with Prof. White (St Andrews) we are interested in understanding DNA repair mechanisms using archaeal organisms as model systems. Some of the proteins we are currently studying include XPB and XPD helicases, endonucleases such as Fen1 and XPF, and the role of the proliferating cell nuclear antigen (PCNA) as a recruiting platform for some of these proteins. We are also investigating the nucleation and filament growth mechanism of archaeal single-strand binding proteins.

 

Single-molecule studies of light-emitting polymers in solution

 

Conjugated polymers are very promising for organic light-emitting diode (OLED) displays as well as solar cells and transistors. The display of information - on mobile phones, televisions and monitors is very important for work, communication, entertainment and learning. Advances in display technology have been dramatic and some of the most attractive are OLEDs. To meet the growing technological opportunities, it is crucial to develop a deeper understanding of how the properties of conjugated polymers relate to their conformation. The conformation is the shape of the polymer and it has a huge effect on the optical properties but it is very difficult to study because every polymer chain has a different shape. We will overcome this problem by measuring single polymers in solution, and by developing techniques to mechanically manipulate the conformation of the polymer to identify how changes in shape alter its light emission properties. This is a pioneering area because single-molecule methods for non-aqueous environments are still in their infancy and our work will allow us to understand how the solvent affects polymer conformation and light emission with an unprecedented level of detail. This project is in collaboration with Prof Samuel (St Andrews), Prof Sakabara (Strathclyde) and Prof Bazan (Univ California Santa Barbara).