The ribosome comes to life.

The ribosome, together with its tRNA substrates, links genotype to phenotype by translating the genetic information carried by mRNA into protein. During the past half-century, the structure and mechanisms of action of the ribosome have emerged from mystery and confusion. It is now evident that the ribosome is an ancient RNA-based molecular machine of staggering structural complexity and that it is fundamentally similar in all living organisms.

Implication of nucleotides near the 3' end of 16S rRNA in guarding the translational reading frame.

Loss of the translational reading frame leads to misincorporation and premature termination, which can have lethal consequences. Based on structural evidence that A1503 of 16S rRNA intercalates between specific mRNA bases, we tested the possibility that it plays a role in maintenance of the reading frame by constructing ribosomes with an abasic nucleotide at position 1503. This was done by specific cleavage of 16S rRNA at position 1493 using the colicin E3 endonuclease and replacing the resulting 3'-terminal 49mer fragment with a synthetic oligonucleotide containing the abasic site using a novel splinted RNA ligation method.

The role of GTP hydrolysis by EF-G in ribosomal translocation.

Translocation of transfer RNA (tRNA) and messenger RNA (mRNA) through the ribosome is catalyzed by the GTPase elongation factor G (EF-G) in bacteria. Although guanosine-5'-triphosphate (GTP) hydrolysis accelerates translocation and is required for dissociation of EF-G, its fundamental role remains unclear. Here, we used ensemble Förster resonance energy transfer (FRET) to monitor how inhibition of GTP hydrolysis impacts the structural dynamics of the ribosome.

The universally conserved nucleotides of the small subunit ribosomal RNAs.

The ribosomal RNAs, along with their substrates the transfer RNAs, contain the most highly conserved nucleotides in all of biology. We have assembled a database containing structure-based alignments of sequences of the small-subunit rRNAs from organisms that span the entire phylogenetic spectrum, to identify the nucleotides that are universally conserved. In its simplest (bacterial and archaeal) forms, the small-subunit rRNA has ∼1500 nt, of which we identify 140 that are absolutely invariant among the 1961 species in our alignment.