Prediction of Charge-Induced Molecular Alignment of Biomolecules Dissolved in Dilute Liquid-Crystalline Phases

Abstract

Alignment of macromolecules in nearly neutral aqueous lyotropic liquid-crystalline media such as bicelles, commonly used in macromolecular NMR studies, can be predicted accurately by a steric obstruction model (Zweckstetter and Bax, 2000). A simple extension of this model is described that results in improved predictions for both the alignment orientation and magnitude of protein and DNA solutes in charged nematic media, such as the widely used medium of filamentous phage Pf1. The extended model approximates the electrostatic interaction between a solute and an ordered phage particle as that between the solute's surface charges and the electric field of the phage. The model is evaluated for four different proteins and a DNA oligomer. Results indicate that alignment in charged nematic media is a function not only of the solute's shape, but also of its electric multipole moments of net charge, dipole, and quadrupole. The relative importance of these terms varies greatly from one macromolecule to another, and evaluation of the experimental data indicates that these terms scale differently with ionic strength. For several of the proteins, the calculated alignment is sensitive to the precise position of the charged groups on the protein surface. This suggests that NMR alignment measurements can potentially be used to probe protein electrostatics. Inclusion of electrostatic interactions in addition to steric effects makes the extended model applicable to all liquid crystals used in biological NMR to date.

Structural basis for RNA recognition by a type II poly(A)-binding protein

“…XlePABP2-TRP bound the A 16 and A 30 RNAs (Fig. 1D) (16) yielded an excellent correlation coefficient (R ϭ 0.99) with one of the three principal axes of the alignment tensor coincident with the 2-fold symmetry axis of the homodimer. In support of the NMR analysis, gel filtration chromatography indicated that the XlePABP2-TRP protein eluted as a single peak with a retention volume that indicated a molecular mass of 28 kDa, approximately twice its predicted size of 16 kDa (Fig.…”

“…A recombinant version of the protease-resistant domain (XlePABP2-TRP) was compared with the intact XlePABP2 protein in binding assays by using poly(A) RNAs (13). XlePABP2-TRP bound the A 16 and A 30 RNAs (Fig. 1D) (16) yielded an excellent correlation coefficient (R ϭ 0.99) with one of the three principal axes of the alignment tensor coincident with the 2-fold symmetry axis of the homodimer.…”

Multiple Alignment Tensors from a Denatured Protein

“…However, the measurement of RDCs corresponding to an additional two independent alignments now provides a more refined view, indicating that a specific tertiary structural arrangement, not inconsistent with a nativelike topology, also persists in the denatured state. Since the mechanisms of electrostatic alignment require an asymmetric distribution of ionizable groups, either on the surface where they mediate transient binding or globally as described by a multipole expansion, 13 these data demonstrate an asymmetry in the distribution of charged residues, in addition to an asymmetry in the distribution of bond vectors. From the observed ensembleaveraged distribution of backbone orientations, one can view a denatured protein as spanning three-dimensional space with a large heterogeneous distribution of conformations yet unable to access all possible segment orientations.…”

A Structural Analysis of DNA Binding by Myelin Transcription Factor 1 Double Zinc Fingers

“…Alignment was assessed by measuring the D 2 O splitting (19 Hz). The program PALES (20) was used for the calculation of the magnitude and orientation of the sterically induced alignment tensor (see below for details). NMR data were processed using Topspin (Bruker, Karlsruhe) and analyzed with SPARKY 3 (37).…”