Molecular dynamics simulations of sarcin-ricin rRNA motif

Abstract

Explicit solvent molecular dynamics (MD) simulations were carried out for sarcin-ricin domain (SRD) motifs from 23S (Escherichia coli) and 28S (rat) rRNAs. The SRD motif consists of GAGA tetraloop, G-bulged cross-strand A-stack, flexible region and duplex part. Detailed analysis of the overall dynamics, base pairing, hydration, cation binding and other SRD features is presented. The SRD is surprisingly static in multiple 25 ns long simulations and lacks any non-local motions, with root mean square deviation (r.m.s.d.) values between averaged MD and high-resolution X-ray structures of 1-1.4 A. Modest dynamics is observed in the tetraloop, namely, rotation of adenine in its apex and subtle reversible shift of the tetraloop with respect to the adjacent base pair. The deformed flexible region in low-resolution rat X-ray structure is repaired by simulations. The simulations reveal few backbone flips, which do not affect positions of bases and do not indicate a force field imbalance. Non-Watson-Crick base pairs are rigid and mediated by long-residency water molecules while there are several modest cation-binding sites around SRD. In summary, SRD is an unusually stiff rRNA building block. Its intrinsic structural and dynamical signatures seen in simulations are strikingly distinct from other rRNA motifs such as Loop E and Kink-turns.

Figure 1

(a) The studied SRD motifs. Base pairs are labeled according to Leontis et al. (3). Original numbering is used in this study. (b) Stereo view of the SRD motif (ECOLI). The S-turn is visible at the fore.

Figure 2

Non-Watson–Crick base pairs in crystal structures of studied SRD motifs. Tetraloop region, (a) trans SE/H G/A base pair; G-bulged region, (b) G/U/A triplet of cis SE/H G/U and trans WC/H U/A base pairs; flexible region, (c) trans H water-mediated A/C base pair; (d) trans SE/H water-mediated U/C base pair, (e) trans H A/A base pair and (f) trans SE/H C/C base pair. (a–d) are based on the crystal structure of ECOLI system (PDB, 1Q9A), (e and f) are based on the crystal structure of mutated RAT system (PDB, 1Q96). (g) Stereo view of the flexible region of the RAT crystal structure (PDB, 430D) with unpaired bases in A/A and C/C base pairs (highlighted).

Figure 3

Different geometries of U/C base pair observed in MD simulations.

Figure 4

Reorientation of A2660 base (highlighted). Original anti conformation is in the upper left corner, the alternative syn conformation in the upper right corner, intermediates in the base-flipping process are below.

Figure 5

Backbone switches observed in MD simulations. Switching phosphates are presented as orange balls. Labeling of phosphates in magenta circles is consistent with presented stereo views: G2663 switch (A); U2656 switch (B); and G2668 switch (C). Crystal structure is in cyan, the averaged MD structure in magenta, the backbone around the switched phosphate in bold.

Figure 6

Hydration and ion-binding sites observed in MD simulations. Ions and water molecules (only oxygens are shown) are black and orange balls, respectively. (A) Tetraloop area—ion positioned near N7(G2659), waters at IIa, IIb and IId positions, (B) ECOLI flexible region—ions near O4(U2653) and N7(G2668); waters at IV, IVac, IVuc and V sites.

Figure 7

Water-mediated tetraloop G/A base pair in MD simulations. Water molecule interconnects N3(G) and N6,N7(A) atoms.