Representation of the superhelical pitches and changes of the distances in the DNA-free and DNA-bound systems.

<p>(A) The cartoon representation of 11.5 TAL repeats and the DNA. The superhelical pitch of dHax3 is assessed by the distance between the Cα atoms of G303 (Gly303) and G675 (Gly675) (left), and the pitch of DNA by the distance between the C3′ atoms of 1dC and 12dT (right). (B) The distance change versus simulation time (solid line) and the value from crystal structure (dotted line) in the DNA-free system. (C) The distance change versus simulation time (solid line) and the value from crystal structure (dotted line) in the DNA-bound system.</p

TALE-DNA direct hydrogen bonds in 11.5 repeats with occupancy over 40%.

<p>Residue id* is the index of a residue in each TAL repeat sequence. The specific interactions are in bold while non-bold denotes nonspecific interactions.</p

Exploring the molecular basis of RNA recognition by the dimeric RNA-binding protein via molecular simulation methods

<p>RNA-binding protein with multiple splicing (RBPMS) is critical for axon guidance, smooth muscle plasticity, and regulation of cancer cell proliferation and migration. Recently, different states of the RNA-recognition motif (RRM) of RBPMS, one in its free form and another in complex with CAC-containing RNA, were determined by X-ray crystallography. In this article, the free RRM domain, its wild type complex and 2 mutant complex systems are studied by molecular dynamics (MD) simulations. Through comparison of free RRM domain and complex systems, it's found that the RNA binding facilitates stabilizing the RNA-binding interface of RRM domain, especially the C-terminal loop. Although both R38Q and T103A/K104A mutations reduce the binding affinity of RRM domain and RNA, the underlining mechanisms are different. Principal component analysis (PCA) and Molecular mechanics Poisson-Boltzmann surface area (MM/PBSA) methods were used to explore the dynamical and recognition mechanisms of RRM domain and RNA. R38Q mutation is positioned on the homodimerization interface and mainly induces the large fluctuations of RRM domains. This mutation does not directly act on the RNA-binding interface, but some interfacial hydrogen bonds are weakened. In contrast, T103A/K104A mutations are located on the RNA-binding interface of RRM domain. These mutations obviously break most of high occupancy hydrogen bonds in the RNA-binding interface. Meanwhile, the key interfacial residues lose their favorable energy contributions upon RNA binding. The ranking of calculated binding energies in 3 complex systems is well consistent with that of experimental binding affinities. These results will be helpful in understanding the RNA recognition mechanisms of RRM domain.</p

Distances between the Cα of G13 and the 5-methyl group of thymine in all repeats with RVDs NG.

<p>All distances are measured in the unit of Å.</p>a<p>D_Initial is the distance from the crystal structure.</p>b<p>D_Average is the mean value of distances from the equilibrium trajectory.</p>c<p>ΔD describes the deviation between D_Initial and D_Average.</p

Structures and domain organization (PDB codes: 3V6P and 3V6T).

<p>(A) The structures of dHax3 in the DNA-free state (left) and DNA-bound state (right). Each structure contains an 11.5-repeat domain, forming a right-handed superhelical assembly. The 11.5 repeats are colored separately. (B) The 11.5-repeat domain mediates DNA binding. Each repeat recognizes one specific nucleotide by using the RVD residues at positions 12 and 13. Single-letter abbreviations for the amino acid residues are as follows: A, Ala; C, Cys; D, Asp; E, Glu; F, Phe; G, Gly; H, His; I, Ile; K, Lys; L, Leu; M, Met; N, Asn; P, Pro; Q, Gln; R, Arg; S, Ser; T, Thr; V, Val; W, Trp; and Y, Tyr.</p

Changes of axis bend angles along the DNA target sequence.

<p>(A) Comparison of the average values (pink) of the axis bend calculated from the equilibrium trajectory along the d(CCCTTTATCTCT) with the corresponding crystal values (sky blue). (B) Fluctuations of axis bend for all dinucleotide steps along the d(CCCTTTATCTCT) during 20 ns trajectory. The color bar gives the variations in bend from 0° (dark blue) to 10° (dark red).</p

Comparison of the average values (pink) of groove widths calculated from the equilibrium trajectory along the d(CCCTTTATCTCT) with the corresponding crystal values (sky blue).

<p>(A) Minor groove widths. (B) Major groove widths. The widening of the major groove is more remarkable at the sites of C2 and C9.</p

Comparison of the average values (pink) of base pair step parameters calculated from the equilibrium trajectory along the d(CCCTTTATCTCT) with the corresponding crystal values (sky blue).

<p>(A) Slide. (B) Roll angle. (C) Twist angle. (D) Rise.</p

Water-mediated hydrogen bonds in 11.5 repeats with occupancy over 40%.

<p>Residue id* is the index of a residue in each TAL repeat sequence.</p

Comparative MD analysis of DNA-free dHax3 (yellow) and DNA-bound dHax3 (dHax3: sky blue; DNA: orange) systems.

<p>(A) The RMSD values of the dHax3 and DNA backbone atoms versus simulation time. (B) The probability distribution of RMSD calculated from the equilibrium trajectories. (C) The RMSF values of the dHax3 Cα atoms calculated from the equilibrium trajectories. (D) The cartoon representation of 11.5 TAL repeats for the dHax3 in the DNA-bound system. The residues in red have relatively higher RMSF values (>1.8 Å) while the ones in blue have relatively lower RMSF values (<1 Å). The other regions are colored white.</p