Anomalous small angle x-ray scattering simulations: proof of concept for distance measurements for nanoparticle-labelled biomacromolecules in solution.

Anomalous small angle X-ray scattering can in principle be used to determine distances between metal label species on biological molecules. Previous experimental studies in the past were unable to distinguish the label-label scattering contribution from that of the molecule, because of the use of atomic labels; these labels contribute only a small proportion of the total scattering signal. However, with the development of nanocrystal labels (of 50-100 atoms) there is the possibility for a renewed attempt at applying anomalous small angle X-ray scattering for distance measurement. This is because the contribution to the scattered signal is necessarily considerably stronger than for atomic labels. Here we demonstrate through simulations, the feasibility of the technique to determine the end-to-end distances of labelled nucleic acid molecules as well as other internal distances mimicking a labelled DNA binding protein if the labels are dissimilar metal nanocrystals. Of crucial importance is the ratio of mass of the nanocrystals to that of the labelled macromolecule, as well as the level of statistical errors in the scattering intensity measurements. The mathematics behind the distance determination process is presented, along with a fitting routine than incorporates maximum entropy regularisation

A New Polarized Target for Neutron-scattering Studies On Biomolecules - 1st Results From Apoferritin and the Deuterated 50s Subunit of Ribosomes

A new polarized target for neutron scattering has been designed by CERN and tested successfully using the reactor FRG-1 at the GKSS Research Centre. The nuclear spins are aligned with respect to the external field - parallel or antiparallel - by dynamic nuclear polarization (DNP). To avoid absorption of neutrons by He-3 the frozen solutions of biomolecules are immersed in liquid He-4 which in turn is thermally coupled to the cooling mixture of He-3/He-4 of the dilution refrigerator. Compared with earlier experiments where the sample had been cooled directly by He-3, the rate of detectable neutrons increased by a factor of 30. Another factor of 30 is due to the installation of the cold source and the beryllium reflector in FRG-1. Polarized neutron scattering from apoferritin in deuterated solvent shows that the proton spin polarization is homogeneous in apoferritin molecules. After saturation of proton nuclear magnetic resonance (NMR), polarized neutron scattering is dominated by deuteron spin contrast. With the deuterated large subunit of E. coli ribosomes, three different basic scattering functions are derived from spin-contrast variation, reflecting the known scattering-length-density distribution of the architecture of rRNA and ribosomal proteins. The planned in situ structure determination of a mRNA fragment is discussed in the light of the present results

Structural Analysis of Stained and Unstained Two-Dimensional Ribosome Crystals

The Italian lizaid, Lacerta sicula, develops large ery stalline sheets of ribosomes during hibernation. The sheets form on the endoplasmic reticulum membrane of previtellogenic oocytes and tend to aggregate into Clusters measuring 10–20 μm across [2]. Thin sections through the clusters show each sheet to be composed of two layers of ribosomes. These double layers are lined on either side with endoplasmic reticulum membranes (Fig. 1a). Sections cut through sheets at an oblique angle show that each layer represents a two-dimensional crystal of ribosome tetramers arranged on a square lattice (Fig. 1b). The space group of the two-dimensional crystals is P4, the until cell dimensions is 595 Å. Both layers have the same symmetry but face in opposite directions so that they appear to b of opposite hand in sections. It can be shown that the layer against the membrane always has the configuration shown in Fig. 2 when seen from above (from here on referred to as “right-handed” configuratio