Sequence-specific size, structure, and stability of tight protein knots

Abstract

Approximately 1% of the known protein structures display knotted configurations in their native fold but their function is not understood. It has been speculated that the entanglement may inhibit mechanical protein unfolding or transport, e.g., as in cellular threading or translocation processes through narrow biological pores. Here we investigate tigh peptide knot (TPK) characteristics in detail by pulling selected 3_1 and 4_1-knotted peptides using all-atom molecular dynamics computer simulations. We find that the 3_1 and 4_1-TPK lengths are typically Delta l~4.7 nm and 6.9 nm, respectively, for a wide range of tensions (F < 1.5 nN), pointing to a pore diameter of ~2 nm below which a translocated knotted protein might get stuck. The 4_1-knot length is in agreement with recent AFM pulling experiments. Detailed TPK characteristics however, may be sequence-specific: we find a different size and structural behavior in polyglycines, and, strikingly, a strong hydrogen bonding and water molecule trapping capability of hydrophobic TPKs due to side chain shielding of the polar TPK core. Water capturing and release is found to be controlla ble by the tightening force in a few cases. These mechanisms result into a sequence-specific 'locking' and metastability of TPKs what might lead to a blocking of knotted peptide transport at designated sequence-positions. Intriguingly, macroscopic tight 4_1-knot structures are reproduced microscopically ('figure-of-eight' vs. the 'pretzel') and can be tuned by sequence in contrast to mathematical predictions. Our findings may explain a function of knots in native proteins, challenge previous studies on macromolecular knots, and may find use in bio- and nanotechnology.