Controlling the nanoparticle's (NP) ability to facilitate cell uptake is particularly desirable for biomedical applications, such as drug delivery. Despite growing evidence highlighting the pivotal role of mechanical properties in biological performances of NPs, consensus regarding how stiffness regulates the cell entry of NPs is lacking. Here, we design and elucidate cell membrane wrapping on soft liposomes in which rigid bowl-like nanostructures are embedded. Compared with pure liposomes, which are often partially wrapped by a membrane due to the high energy barrier associated with the oblate deformation, embedded nanobowls provide physical support to promote wrapping by increasing the liposome rigidity. Through correlating liposome deformation, invagination, and nanobowl orientation, two distinct pathways of cell membrane wrapping on nanobowl-supported liposomes are identified. Different from alternative methods of tuning the rigidity of NPs uniformly, the nanobowl-supported liposome is characterized by mechanical heterogeneity and nanobowl rotation, which correlates with liposome deformation to promote endocytic wrapping. Once the nanobowl's rotation is restrained, wrapping turns out to be accomplished via membrane protrusion rather than invagination, and the wrapped liposome could be trapped with frustrated internalization and potential membrane perturbation. Following this principle, moderate decreases of the nanobowl size and adhesive strength with the liposome can facilitate the nanobowl's rotation to promote cell uptake. These results are expected to improve our ability to develop better NPs for enhanced drug delivery.

The knot is one of the most remarkable topological features identified in an increasing number of proteins with important functions. However, little is known about how the knot is formed during protein folding, and untied or maintained in protein unfolding. By means of all-atom molecular dynamics simulation, here we employ methyltransferase YbeA as the knotted protein model to analyze changes of the knotted conformation coupled with protein unfolding under thermal and mechanical denaturing conditions. Our results show that the trefoil knot in YbeA is occasionally untied via knot loosening rather than sliding under enhanced thermal fluctuations. Through correlating protein unfolding with changes in the knot position and size, several aspects of barriers that jointly suppress knot untying are revealed. In particular, protein unfolding is always prior to knot untying and starts preferentially from separation of two α-helices (α1 and α5), which protect the hydrophobic core consisting of β-sheets (β1–β4) from exposure to water. These β-sheets form a loop through which α5 is threaded to form the knot. Hydrophobic and hydrogen bonding interactions inside the core stabilize the loop against loosening. In addition, residues at N-terminal of α5 define a rigid turning to impede α5 from sliding out of the loop. Site mutations are designed to specifically eliminate these barriers, and easier knot untying is achieved under the same denaturing conditions. These results provide new molecular level insights into the folding/unfolding of knotted proteins.

No abstract