Probing Mechanical Properties of Biomolecules using Nanopores

Nanopore translocation is a promising label-free single molecule technique to distinguish between bio-molecules. The confined nature of the nanopore restricts the allowed conformations of the molecules or necessitates a conformational change. These effects are reflected in the observed current traces and thus help us in measuring the flexibility of these biomolecules. Recently, we applied molecular dynamics simulations using a structure-based model to observe a correlation between the maximum RMSF of the protein and the width of the experimental current blockade distribution. This suggests that protein translocation can be utilized as a high-throughput method to distinguish between functional conformers in proteins. Applying this technique to translocation of tRNA offers an interesting challenge since the tRNA is expected to undergo a conformational change due to the constricted size of the nanopore. Using the same energy landscape techniques, we have calculated the mean first passage time (MFPT) for crossing the rate-limiting free-energy barrier for multiple tRNA species. We find agreement between the MFPT values and the experimental translocation times. Further, these calculations suggest that the experiments specifically observe transient partial unfolding of tRNA.