Structural investigation of the protein-RNA complex directing the incorporation of the amino acid selenocysteine and the RNA that binds the SAM domain of the Smaug protein

Nuclear magnetic resonance (NMR) structural studies were completed on the selenocysteine insertion sequence (SECIS) for Escherichia coli formate dehydrogenase N (fdnG). Structural studies were completed using an 18mer RNA construct and a construct containing a 3' dangling adenine. It was determined that fdnG adopts a hairpin structure containing a bulged uridine residue in the stem and a triloop structure containing stacked nucleotides. Improved NMR spectra acquired in H2O was observed for the construct containing a 3' dangling adenosine. To further investigate the role of 3' dangling nucleotides on NMR signal enhancement, variants of fdnG RNA containing the four possible 3' dangling nucleotides were studied using NMR and compared for NMR signal enhancement. It was determined that 3' adenosine and 3' guanosine give the greatest signal enhancement.;The sterile alpha motif (SAM) domain of Saccharomyces cerevisiae post-transcriptional regulator Vts1p has previously been shown to have high affinity for RNA hairpins containing a CUGGC pentaloop. NMR structural studies were completed on a 19mer RNA construct containing the CUGGC pentaloop. It was determined that this RNA consists of an A-form helical stem, capped with an ordered tetraloop structure.