Search

Stochastic search algorithms can be used to perform rapid six-dimensional molecular-replacement searches. A molecular-replacement procedure has been developed that uses an evolutionary algorithm to simultaneously optimize the orientation and position of a search model in a unit cell. Here, the performance of this algorithm and its dependence on search model quality and choice of target function are examined. Although the evolutionary search procedure is capable of finding solutions with search models that represent only a small fraction of the total scattering matter of the target molecule, the efficiency of the search procedure is highly dependent on the quality of the search model. Polyalanine models frequently provide better search efficiency than all-atom models, even in cases where the side-chain positions are known with high accuracy. Although the success of the search procedure is not highly dependent on the statistic used as the target function, the correlation coefficient between observed and calculated structure-factor amplitudes generally results in better search efficiency than does the R factor. An alternative stochastic search procedure, simulated annealing, provides similar overall performance to evolutionary search. Methods of extending the evolutionary search algorithm to include internal optimization, selection and construction of the search model are now beginning to be investigated. [ABSTRACT FROM AUTHOR]

The hepatitis C (HCV) internal ribosome entry site (IRES) element plays a central role in cap-independent translation of the viral genomic RNA. The unique conformation of IRES domain II is critical for 80S ribosomal assembly and initiation of viral translation. Here, the crystal structure of subdomain IIa of the HCV IRES has been determined at 2.3 A resolution, revealing the positions of divalent metal ions and complex inter-strand interactions that stabilize the L-shaped conformation of the RNA. The presence of divalent metal ions was necessary for crystal formation. Magnesium ions occupy specific sites that appear to be critical for the formation of the folded conformation. Subdomain IIa also was crystallized in the presence of strontium, which improved the diffraction quality of the crystals and the ability to identify interactions of the RNA with metal ions and tightly bound water molecules. The hinge region and noncanonical G-U base-pair motifs are stabilized by divalent metal ions and provide unique structural features that are potential interaction sites for small-molecule ligands. The information obtained from the crystal structure provides a basis for structure-guided design of HCV translation inhibitors targeting disruption of ribosomal assembly.