The objective of this project is the determination of the structures of biologically relevant RNAs and ribonucleoprotein complexes by NMR. The information it generates will lead to a better understanding of the chemical basis of the biological processes in which these RNAs participate, and will contribute to the store of information available about the conformational properties of RNA, which are still poorly understood. Since RNA plays a critical role in almost every aspect of gene expression, this knowledge is fundamental to understanding many physiological processes in humans, both normal and pathological. The first targets of this study will be: (1) the loop A and helix II-III domains of 5S rRNA from E. coli, (2) ribosomal protein L18 and the complex it forms with 5S rRNA, and (3) the loop E region from spinach chloroplast 5S rRNA. Over the years, no RNA has been studied more intensively by chemical and enzymatic means than that 5S rRNA. In addition to providing a structure for this important ribosomal component, the studies proposed here enable the community to better understand chemical probing data on 5S rRNA, and that should enable them to interpret more accurately data obtained from RNAs less amenable to structure determination. The chloroplast project will cast light on the conformational significance of sequence homology in RNA. Later in the grant period, EBER II RNA and its complex with ribosomal protein L22 will be examined, as will the conformation of the loop IIa/IIb region from yeast U2 snRNA. Running in parallel with these activities will be initiatives aimed at improving the quality of the structures RNA spectroscopists. The most important of these has to do with the measurement and use of residual dipolar coupling data to improve the long-range accuracy of RNA solution structures, which is rapidly developing area of the macromolecu1ar NMR field.
