Refinement of the solution structure of the DNA decamer 5'd(CTGGATCCAG)2: combined use of nuclear magnetic resonance and restrained molecular dynamics

Abstract

The solution structure of the self-complementary DNA decamer 5'd(CTGGATCCAG)2 comprising the specific target site for the restriction endonuclease BamH1 is investigated by using nuclear magnetic resonance sectroscopy and restrained molecular dynamics. With the exception of the H5'/H5" sugar proton resonances, all the nonexchangeable proton resonances are assigned sequentially by using pure-phase absorption two-dimensional nuclear Overhauser enhancement spectroscopy. From the time dependence of the nuclear Overhauser effects a set of 160 approximate interproton distances is determined and used as the basis of a structure refinement employing restrained molecular dynamics in which the interproton distances are incorporated into the total energy function of the system in the form of an effective potential term. Two restrained dynamics simulations are carried out, starting from classical B- and A-DNA [atomic root mean square (rms) difference 5.7 A]. In both cases convergence is achieved to very similar B-type structures with an atomic rms difference of 0.9 A which is comparable to the rms fluctuations of the atoms about their average positions. In addition, the rms difference between the experimental and calculated values of the interproton distances for both average restrained dynamics structures is approximately 0.3 A. These results suggest that the converged restrained molecular dynamics structures represent reasonable approximations of the solution structure. The average restrained dynamics structures exhibit clear sequence-dependent variations of torsion angles and helical parameters. In addition, the structures exhibit a small bend of around 10-20 degrees at the second (TpG) and eighth (CpA) base pair steps. This can be attributed to the positive base roll angles and large base pair slide values at the two Pyr-Pur steps. The central core of the decamer comprising the six-base recognition site for BamH1 (GGATCC), however, is straight.

Structure determination of [d(ATATATAUAT)]2 via two‐dimensional NOE spectroscopy and molecular dynamics calculations

“…There is partial interstrand overlap of the six-member rings of the adenines in the T-A step, which results in a smaller helical twist angle. This pattern has been seen by others (Yoon et al, 1988;Nilges et al, 1987) for alternating purine-pyrimidine sequences, with the value of the helical twist being lower for the purine-pyrimidine step.…”

“…Twodimensional (2D) NMR, in particular 2D NOE spectra, when used in conjunction with energy-refinement calculations, can be used to determine the high-resolution structure of DNA fragments where subtle changes in structure can be detected (Zhou et al, 1987;Nilges et al, 1987;Nikonowicz et al, 1989;Pieters et al, 1990). A major obstacle to accurately depicting a structure using this methodology is the problem of extracting a large number of accurate interproton distances from the 2D NOE cross-peak intensities.…”

TWo-Dimensional1H and31P NMR Spectra and Restrained Molecular Dynamics Structure of an Oligodeoxyribonucleotide Duplex Refined via a Hybrid Relaxation Matrix Procedure

“…As shown by these and other results (19,64) NOESY distances appear to be able to restrain the calculated structures to conformations that accurately reflect these sequence-specific variations in the local conformation of the DNA. Even distances derived from the two-spin approximation treatment of the NOESY data are accurate enough to reproduce these large local variations in structure.…”

“…The correlation between calculated and predicted values are very poor (correlation coefficient between the Calladine rules and AMBER calculated helix twist values is a negative 0.48). However, NOESY distance-restrained molecular dynamics calculations generally show much better correlation of predicted and observed helical twist values (19,64; see below). Molecular dynamics calculations should be better able to overcome small energy barriers (on the order of kT) that otherwise limit the ability of an energy minimization scheme to locate a global energy minimum.…”

“…The development of sequence-specific, two-dimensional nuclear magnetic resonance (2D NMR) assignment methodologies (5)(6)(7)(8)(9)(10)(11)(12)(13)(14) and higher-field spectrometers have made the study of modest size oligonucleotides (10-20 base pairs) possible. Two-dimensional NMR, combined with distance geometry (15)(16)(17) or restrained molecular mechanics/dynamics (3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20), is now capable of elucidating the fine structure of short DNA duplexes in solution. Unfortunately, evaluation of interproton distances from a 2D-NMR nuclear Overhauser effect spectroscopy (NOESY) experiment has relied on the so-called "two-spin approximation" (17,21).…”