Mechanical Stabilization of Proteins due to Ligand Binding using Magnetic Tweezers
Mentor 1
Ionel Popa
Mentor 2
Narayan Dahal
Start Date
1-5-2020 12:00 AM
Description
The mechanical unfolding of proteins is commonly measured using single molecule force spectroscopy techniques. Here, we use magnetic tweezers and hetero-covalent attachments to measure this unfolding with the ability to change the surrounding solution or solvent easily in order to assess binding of a ligand to its substrate. Protein L and the Talin rod domains, R7 and R8, are our two substrates of choice. Protein L is a bacterial protein with two binding sites for kappa-light chain antibody ligands. The R8 domain of Talin is known to bind a potential antitumor ligand, Deleted in Liver Cancer 1 (DLC1). HaloTag and Spytag were used on the ends of each molecular construct to tether the proteins. Magnetic tweezers are then able to expose the molecule to forces up to 100 pN and measure unfolding and refolding cycles. We find that ligand binding induces mechanical stabilization in both protein L and Talin R8 domains. These results open the way for screening new mechano-active drugs against bacterial infection and cancer.
Mechanical Stabilization of Proteins due to Ligand Binding using Magnetic Tweezers
The mechanical unfolding of proteins is commonly measured using single molecule force spectroscopy techniques. Here, we use magnetic tweezers and hetero-covalent attachments to measure this unfolding with the ability to change the surrounding solution or solvent easily in order to assess binding of a ligand to its substrate. Protein L and the Talin rod domains, R7 and R8, are our two substrates of choice. Protein L is a bacterial protein with two binding sites for kappa-light chain antibody ligands. The R8 domain of Talin is known to bind a potential antitumor ligand, Deleted in Liver Cancer 1 (DLC1). HaloTag and Spytag were used on the ends of each molecular construct to tether the proteins. Magnetic tweezers are then able to expose the molecule to forces up to 100 pN and measure unfolding and refolding cycles. We find that ligand binding induces mechanical stabilization in both protein L and Talin R8 domains. These results open the way for screening new mechano-active drugs against bacterial infection and cancer.
