The biogenesis and function of mature microRNAs (miRNAs) is dependent on the nuclear export of miRNA precursors (premiRNA) by . To characterize the molecular mechanisms of how pre-miRNA is recognized and transported by Exp5, we have performed 21 molecular dynamic (MD) simulations of RNA-bound Exp5 (Exp5-RanGTP-premiRNA, Exp5-RanGDP-premiRNA, Exp5-premiRNA), RNA-unbound Exp5 (Exp5-RanGTP, Exp5-RanGDP, apo-Exp5), and pre-miRNA. Our simulations with standard MD, steered molecular dynamics (SMD), and energy analysis have shown that (1) Free Exp5 undergoes extensive opening motion, and in this way facilitates the RanGTP binding. (2) RanGTP efficiently regulates the association/dissociation of pre-miRNA to its complex by inducing conformational changes in the HEAT-repeat helix stacking of Exp5. (3) The GTP hydrolysis prevents Ran from rebinding to Exp5 by regulating the hydrophobic interfaces and salt bridges between Ran and Exp5. (4) The transition from the A9-form to the A-form of the pre-miRNA modulates the structural complementarities between the protein and the pre-miRNA, thus promoting efficient assembly of the complex. (5) The baseflipping process (from the closed to the fully flipped state) of the 2-nt 39 overhang is a prerequisite for the pre-miRNA recognition by Exp5, which occurs in a sequence-independent manner as evidenced by the fact that different 2-nt 39 overhangs bind to Exp5 in essentially the same way. And finally, a plausible mechanism of the pre-miRNA export cycle has been proposed explaining how the protein-protein and protein-RNA interactions are coordinated in physiological conditions.

Wrapping of single-wall carbon nanotubes (SWCNTs) by singlestranded DNA (ssDNA) was found to be sequence-dependent, offering properties such as the facilitation of SWCN sorting, ultrafast DNA sequencing, and construction of chemical sensors. Although the interactions of nucleic acids with SWCNTs have been studied thoroughly, the DNAÀCNT hybrid especially for the oligonucleotides containing more than one nucleotide has not yet been fully understood.To address this, we have examined new and unconventional DNA dinucleotides involving all 16 combinations of two DNA nucleotides attached with chiral (8,4) and armchair (6,6) SWCNTs using all-atom molecular dynamics simulations and thermodynamic analyses. The 16 dinucleotides with different sequence compositions are found to readily adsorb onto SWCNTs and display interesting binding behaviors such as base flipping, local dynamic stability of structure, and conformational shifting. Four dinucleotides, i.e., AC, AG, GC, and GT, share similar dynamic properties (base turning and conformation transformation) in (8,4) and (6,6) systems. The different dynamic profiles between the compositional isomers with the reverse sequences such as the AG and GA show that the sequence order also impacts the dynamic recognition and binding energy of the ssDNAÀCNT hybrid. Clustering-analysis-derived representative conformations imply that general dinucleotides are inclined to spread on the SWCNT surface, and the adjacent bases tend to stretch away from each other. Dinucleotides like AC, AT, CG, CT, GC, GG, TA, TC, TG, and TT adopt similar geometries on both CNTs, suggesting that their structures are not predominantly influenced by the nanotube chirality but controlled by the identity of the base sequence, sequence order, and the basic cylindric structure of SWCNT. In addition, the nucleotide bases have a high degree of orientational order on the nanotube surface and the orientations of each base are significantly affected by the sequence of DNA and the chirality of nanotube, emphasizing that the structural order plays an important role in the binding of DNA and CNTs. However, our energy analysis shows that due to small different curvatures of the CNT surface, the binding affinity of most dinucleotides (except AG, CA, CG, and TG) to the chiral and armchair nanotube is not significantly different. Generally, the dinucleotides constituted with purine and thymine exhibit the lowest binding free energy, resulting from the van der Waals interactions and solvent effects. The thymine-based dinucleotides reduce the solvation free energy of the SWCNT in aqueous solution more effectively as compared to other bases. The present work also demonstrates that the total binding free energy is sequence specific but not merely a sum of individual baseÀSWCNT binding free energies.

Aurora A is an important oncogenic kinase for mitotic spindle assembly and a potentially attractive target for human cancers. Its activation could be regulated by ATP cycle and its activator TPX2. To understand the activation mechanism of Aurora A, a series of 20 ns molecular dynamics (MD) simulations were performed on both the wild-type kinase and its mutants. Analyzing the three dynamic trajectories (Aurora A-ATP, Aurora A-ADP, and Aurora A-ADP-TPX2) at the residue level, for the first time we find two TPX2-dependent switches, i.e., switch-1 (Lys-143) and switch-2 (Arg-180), which are tightly associated with Aurora A activation. In the absence of TPX2, Lys-143 exhibits a “closed” state, and becomes hydrogen-bonded to ADP. Once TPX2 binding occurs, switch-1 is forced to “open” the binding site, thus pulling ADP away from Aurora A. Without facilitation of TPX2, switch-2 exits in an “open” conformation which accompanies the outward-flipping movement of P·Thr288 (in an inactive conformation), leading to the crucial phosphothreonine exposed and accessible for deactivation. However, with the binding of TPX2, switch-2 is forced to undergo a “closed” movement, thus capturing P·Thr288 into a buried position and locking its active conformation. Analysis of two Aurora A (K143A and R180A) mutants for the two switches further verifies their functionality and reliability in controlling Aurora activity. Our systems therefore suggest two switches determining Aurora A activation, which are important for the development of aurora kinase inhibitors.

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