University of Wisconsin–Madison

Hybrid Methods

NMR and SAXS Hybrid Methods

The global structural information about molecular shapes provided by small-angle X-ray scattering (SAXS) is highly complementary to the high-resolution structural information that can be obtained by NMR. One of the major challenges of studying biomolecules by NMR is that they become difficult to study as their molecular mass increases. SAXS can be used to help overcome the size limitations of NMR because it is not so limited by the size of systems that can be easily studied. Further, SAXS is highly complimentary to NMR given that samples can be studied under the same temperature and buffer conditions. Thus, the atomic resolution information obtained from NMR can be combined with the low-resolution shape information from SAXS to generate a more complete insight into the structures of biomolecules.

transform to 1d opt

The NMRFAM has a Bruker Nanostar bench top system capable of carrying out SAXS experiments on biomolecules and materials. SAXS experiments require only a small amount of material (40-50 mL of 1-10 mg/mL sample) and a full data set can be collected in a single day at the NMRFAM. The Bruker Nanostar system at the NMRFAM is capable of characterizing molecules up to ~22 nm in maximum end-to-end distance. The Bruker Nanostar system can also be used to carry out temperature dependent studies with a temperature range of -30°C to 100°C. The expert staff at the NMRFAM are capable of assisting users in both data collection and analysis.

rna

One example of an NMR/SAXS hybrid technology developed at the NMRFAM involves determining structures of large RNA molecules [1]. The Butcher group recently solved the solution structure of the 111 nucleotide U2/U6 RNA complex (above) by developing a method that combines NMR structural data (NOE’s and RDC’s) with SAXS. This novel method involves generating a large ensemble of RNA structures based on knowledge of the RNA secondary structure. Models from the large structure pool with the best agreement to the NMR and SAXS data are then selected and further refined against the NMR/SAXS data.

1. Burke JE, Sashital DG, Zuo X, Wang YX, Butcher SE. (2012) Structure of the yeast U2/U6 snRNA Complex. RNA 18(4):673-83.

  • tetraloopRNA Structures by MSA/Pf1 RDCs, NOEs and SAXS

We are developing a productive approach for RNA structure determination that combines SAXS and Nuclear Overhauser Effect (NOE) data with Pf1 and Magnetic Susceptibility Anisotropy (MSA) induced Residual Dipolar Couplings (RDCs) collected at multiple fields.

iapv constr small iapv nmr small

iapv saxs small iapv_rdc.gif

iapv el densit iapv small