A new generation of CCP4 monomer library based on Crystallography Open Database

The use of prior chemical knowledge such as bond lengths, bond angles about constituent blocks of macromolecules and ligands is an essential part of macromolecular crystal structure analysis. One of the reliable sources of such chemical knowledge is small molecule database where small molecule crystal structures have been analysed against high-resolution, high-quality experimental data. Furthermore, vast amount of data in small molecule database provide comprehensive coverage of flexible chemical environment and enable proper statistical analysis to avoid biased representation of those chemical properties. This presentation describes our work on organization of the data from open-access and daily-updated small molecule database, Crystallography Open Database(COD) [1], into a new generation of CCP4 monomer library (Dictionary), a container of prior chemical knowledge [2]. In order to describe specific environment atoms are in, they are classified into different atomic types based on local graphs and some basic chemical properties of atoms. This scheme can be applied to any small molecule databases. The atom types, and values of bond lengths and bond associated with them, are further clustered into a hierarchical tree and an isomorphism-mapping algorithm is implemented to facilitate fast search among a large number of atom types (typically several millions). This also provides a mechanism to derive reliable values for bond lengths and angles of novel ligands. Metal and non-organic atoms are treated differently with organic ones. The original data in COD are curated using several criteria and further statistical analysis on derived values of bond lengths and angles are allow to extract reliable chemical information from such databanks as COD. There are several software tools associated with new dictionary including 1) generate "ideal" bond lengths and angles for unknown ligand; 2) generate starting coordinates to represent one of the conformation of the ligand under consideration.

Low Resolution Refinement with ProSMART, COOT and REFMAC5

Membrane proteins and large assemblies are currently a major focus of molecular biology and molecular medicine. Due to their size and flexibility, these structures may only yield poor quality crystals for which diffraction intensities can be measured to merely mid-low resolution. Nevertheless, these data contain valuable structural information. Here, it will be shown how new features in COOT [1], REFMAC5 [2] and ProSMART [3] can help to exploit low resolution data for model building and refinement, as well as aid model validation. Refinement at low resolution can be stabilised with regularisers, such as jelly-body and external restraints. These allow to routinely obtain good quality models even in cases where only low-resolution data are available (e.g. >3Å). External restraints (available for protein and DNA/RNA) exploit structural prior knowledge, utilising the assertion that local interatomic distances should agree with previous observations. Sources for such prior knowledge include isomorphous and homologous structures, hydrogen bonding patterns, and typical conformations of secondary structure elements. Importantly, global rigidity is not enforced by these restraints - the approach presented allows for dramatic conformational differences between target and reference models. Consequently, restraints may be generated using homologous reference models resolved in different crystal forms. COOT facilitates model building at low resolution by removing degrees of freedom through so-called "backrub rotamers" and torsion angle restraints, as well as providing semi-automatic building options such as model morphing and jiggle fit . Map sharpening and blurring, now available in both COOT and REFMAC5, can be employed to provide further insight regarding the validity of a model, as well as aiding the model building process. General guidelines for the application of these features are provided, along with examples demonstrating their usage.

The Structure of the Yeast Mitochondrial Large Ribosomal Subunit

Mitochondria have specialized ribosomes that have diverged from their bacterial and cytoplasmic counterparts. We have solved the structure of the yeast mitoribosomal large subunit using single-particle electron cryo-microscopy. The resolution of 3.2 Ångstroms enabled a nearly complete atomic model to be built de novo and refined, including 39 proteins, 13 of which are unique to mitochondria, as well as expansion segments of mitoribosomal RNA. The structure reveals a new exit tunnel path and architecture, unique elements of the E site and a putative membrane docking site.