The structures, functions, and evolution of Sm -like archaeal proteins (SmAPs)

Sm proteins were discovered nearly 20 years ago as a group of small antigenic proteins (≈90--120 residues). Since then, an extensive amount of biochemical and genetic data have illuminated the crucial roles of these proteins in forming ribonucleoprotein (RNP) complexes that are used in RNA processing, e.g., spliceosomal removal of introns from pre-mRNAs. Spliceosomes are large macromolecular machines that are comparable to ribosomes in size and complexity, and are composed of uridine-rich small nuclear RNPs (U snRNPs). Various sets of seven different Sm proteins form the cores of most snRNPs. Despite their importance, very little is known about the atomic-resolution structure of snRNPs or their Sm cores. As a first step towards a high-resolution image of snRNPs and their hierarchic assembly, we have determined the crystal structures of archaeal homologs of Sm proteins, which we term Sm-like archaeal proteins (SmAPs).;Beginning with a Pyrobaculum aerophilum (Pae ) structural genomics pilot project, we determined the structure of Pae SmAP1. This structure provided the first direct evidence for a toroid-shaped Sm homoheptamer at the snRNP core, and provided many insights and implications for SmAP evolution and RNA binding in Sm cores. Then, in order to extend these results, we solved the structure of Pae SmAP1 and a heptameric methanobacterial SmAP (Mth SmAP1) bound to uridine-5'-monophosphate (UMP); the uracil bases line the heptamer pore in the Mth ligand-bound structure, and suggest a more specific model for RNA binding than we were able to propose earlier.;Further characterization of the oligomerization and ligand-binding properties of Mth and Pae SmAP1s has allowed us to conclude that: (i) SmAPs form several oligomers besides the archetypal heptamer, including 14-mers and fibrillar polymers; (ii) Mth SmAP1 and Archaeoglobus fulgidus (Afu) SmAP1 recognize uracil bases in a nearly identical manner, suggesting a conserved RNA-binding mode for SmAPs; and (iii) Pae and Mth SmAP1s gel-shift supercoiled DNA, perhaps by nonspecific binding to single-stranded DNA. Our sequence analyses shed light on the evolution of Sm proteins: the SmAP module is a phylogenetically well-conserved domain that probably gave rise to modern (eukaryotic) Sm heteroheptamers via gene duplication and neutral drift. Crystal structure determinations for Pae SmAP2 and SmAP3 proteins are currently in progress, and will deepen our understanding of Sm protein function and evolution.;As part of the same Pae structural genomics project, we solved two structures that are unrelated to the SmAP work: an archaeal homolog of survival protein E (Pae SurEalpha) and a putative Pae Nudix protein. The final two chapters of this dissertation describe these structures and their significance.