How do proteins fold and keep us healthy?
High-resolution protein folding and dynamics in the cell and biomolecular spectroscopy. The Cavagnero group focuses on developing new spectroscopic techniques to study the structure/dynamics of biomolecules at high resolution and the fundamental principles of protein folding and misfolding in the cell. We place emphasis on the folding of nascent proteins emerging from the molecular machine responsible for their biosynthesis, the ribosome. We also study how molecular chaperones affect protein folding.
Our work has several practical implications. First, the large-scale production of soluble proteins relevant to biotechnology and medicine, including protein-based drugs, will be enhanced by exploiting the principles we learn from the study of protein folding in the cell. Second, a better understanding of the role of the Hsp70 molecular chaperone in client-protein binding is leading us to rationally design novel antimicrobial agents targeting the protein folding machinery. Third, the mechanistic principles we are learning, focusing on the balance between protein folding and aggregation in the cell, feed useful principles for the control of deleterious aggregation processes leading to a variety of diseases, including prion-related disorders, Parkinson’s disease, Alzheimer's disease, Huntington's chorea and cancer.
Laser-enhanced NMR spectroscopy. We are developing novel laser-assisted methodologies to enhance the power NMR spectroscopy for the analysis of protein folding. Our efforts include laser- enhanced approaches to boost NMR sensitivity, new radio-frequency pulse schemes to overcome undesired NMR resonance line-broadening due to conformational exchange in the intermediate chemical shift timescale, and methods to site-specifically study the role of water in protein folding. Exciting new developments include laser-driven techniques based on photochemically-enhanced dynamic nuclear polarization (photo-CIDNP) and its application to heteronuclear correlation NMR in 15N and 13C isotopically enriched samples.
Protein folding in vivo: conformation and dynamics of ribosome-bound nascent polypeptides. The earliest stages of a protein's life are crucial for its ability to function in the cell. Our group is pioneering the study of protein folding at the ribosomal exit tunnel by a combination of time-resolved fluorescence, multidimensional NMR and chemical biology approaches. In addition, we are developing novel approaches to study protein folding and translation rates in living cells by single-molecule fluorescence microscopy.
Fundamental questions in protein folding.We are involved in the study of fundamental questions in protein folding, including kinetic trapping events across the folding energy landscape, the role of water and hydrophobic collapse, the role of the protein's C terminus in folding, the thermodynamic balance between protein folding and aggregation, and the prediction of structure from amino acid sequence.
The role of molecular chaperones in protein folding and disease. All the above research directions are pursued in the absence and presence of cotranslationally- active chaperones. We are exploring how molecular chaperones such as the bacterial Hsp70, known as DnaK, interact with their substrate proteins and affect their folding mechanisms. This work is carried out primarily by multidimensional NMR on 15N- and 13C-enriched polypeptide substrates and involves both kinetic and structural analysis. In addition, we are designing novel antimicrobial agents interfering with the binding of the molecular chaperone Hsp70 to its natural cellular-protein substrates.