The Noller Lab
The Noller lab studies the structure and function of the ribosome, the cellular ribonucleoprotein complex responsible for translation of the genetic code, and synthesis of proteins in all organisms. Ribosomes are highly conserved, with functional sites that are nearly identical across the phylogenetic spectrum. One of our main contributions has been to show the functional importance of ribosomal RNA (rRNA) in protein synthesis. We provided early evidence for the participation of rRNA in peptide bond formation, tRNA binding, interaction of ribosomes with translational initiation and elongation factors, binding of antibiotics and ribosomal subunit association. The first sequences of the large 16S and 23S rRNAs were obtained in our lab, and their secondary structures deduced by comparative sequence analysis, in collaboration with Carl Woese. We developed methods for chemical footprinting of RNA, and used them to map the rRNA binding sites of the 30S subunit ribosomal proteins, the A-, P- and E-site tRNAs, translation factors and antibiotics. Chemical footprinting of ribosomes during steps of in vitro translation led to the hybrid-states model for translocation. Development of directed hydroxyl radical probing methods allowed us to position ribosomal proteins, translation factors and tRNAs in three dimensions relative to structural features of rRNA. In the late 1990s, we began to use X-ray crystallography, resulting in the structures of functional complexes of the complete ribosome containing bound mRNA and tRNAs at up to 3.0Å resolution.
We are now trying to understand how the three-dimensional structure of the ribosome defines its functional properties. It is clear that the ribosome is a molecular machine. Among the key questions are, what are the ribosome's moving parts, how do they move, how are these movements coordinated and how do they enable the mechanisms of protein synthesis? To this end, we are using methods to study ribosome movement at the molecular level in real time, and in parallel are attempting to solve structures of functional complexes of the ribosome trapped in intermediate stages of the translation process. In collaboration with the Clegg and Ha groups (Illinois), we have studied ribosomal movement with fluorescence resonance energy transfer (FRET), using both bulk and single-molecule approaches, and in collaboration with the Tinoco and Bustamante laboratories (UC Berkeley) we are using optical tweezers to study ribosome dynamics and to measure molecular forces that occur during ribosome movement. Recently, we have used bulk FRET to observe intersubunit movement of the ribosome and rotational movement of the head of the 30S subunit during translocation of mRNA. Using crystallography, we have solved the structure of a complex of the 70S ribosome bound to the GTPase release factor RF3 in which both the body and head of the 30S subunit are trapped in a rotated state, providing insights into the structural basis of these large-scale molecular movements.
Selected Publications: Guo, Z. and Noller, H.F. 2012. Rotation of the head of the 30S ribosomal subunit during mRNA translocation. Proc Natl Acad Sci USA (in press) Zhou, J., Lancaster, L., Trakhanov, S. and Noller, H.F. 2011. Crystal Structure of Release Factor RF3 Trapped in the GTP state on a rotated conformation of the ribosome. RNA 18:230-240. Qu, X., Wen, J.D., Lancaster, L., Noller, H.F., Bustamante, C., and Tinoco, I., Jr. 2011. The ribosome uses two active mechanisms to unwind messenger RNA during translation. Nature 475: 118-121. Zhu, J., Korostelev, A., Costantino, D.A., Donohue, J.P., Noller, H.F. and Kieft, J.S. 2011. Crystal structures of complexes containing domains from two viral internal ribosome entry site (IRES) RNAs bound to the 70S ribosome. Proc Natl Acad Sci USA 108:1839-1844. Ermolenko, D.N. and Noller, H.F. 2011. mRNA Translocation Occurs During the Second Step of Ribosomal Intersubunit Rotation. Nat Struct Mol Biol 18:457-462. Korostelev, A., Zhu, J., Asahara, H. and Noller, H.F. 2010. Recognition of the amber UAG stop codon by release factor RF1. EMBO J 29:2577-85. Korostelev, A., Laurberg, M., and Noller, H.F. 2009. Multistart simulated annealing refinement of the crystal structure of the 70S ribosome. Proc Natl Acad Sci USA 106:18195-18200. Cornish, P.V., Ermolenko, D.N., Staple, D.W., Hoang, L., Hickerson, R., Noller, H.F. and Ha, T. 2009. Following Movement of the L1 Stalk Between Three Functional States in Single Ribosomes. Proc Natl Acad Sci USA 106:2571-2576. Korostelev, A., Asahara, H., Lancaster, L., Laurberg, M. Hirschi, A., Zhu, J., Trakhanov, S., Scott, W. and Noller, H.F. 2008. Crystal Structure of a Translation Termination Complex Formed with Release Factor RF2. Proc Natl Acad Sci USA 105:19684-19689. Lancaster, L., Lambert, N.J., Maklan, E.J., Horan, L.H and Noller, H.F. 2008. The sarcin-ricin loop of 23S rRNA is essential for assembly of the functional core of the 50S ribosomal subunit. RNA 14:1-14. Martick, M., Horan, L., Noller, H., and Scott, W.G. 2008. A discontinuous hammerhead ribozyme embedded in a mammalian messenger RNA. Nature 454(14):899-903. Laurberg, M., Asahara, H., Korostelev, A., Zhu, J., Trakhanov, S. and Noller, H.F. 2008. Structural basis for translation termination on the 70S ribosome. Nature 454(14):852-857. Wen, J.D., Lancaster, L., Hodges, C., Zeri, A.C., Yoshimura, S.H., Noller, H.F., Bustamante, C., and Tinoco, I. 2008. Following translation by single ribosomes one codon at a time. Nature 452(7187):598-603. Cornish, P.V., Ermolenko, D.N., Noller, H.F., and Ha, T. 2008. Spontaneous intersubunit rotation in single riboomes. Mol Cell 30(5):578-88. Ermolenko, D.N., Majumdar, Z.K., Hickerson, R.P., Spiegel, P.C., Clegg, R.M., and Noller, H.F. 2007. Observation of intersubunit movement of the ribosome in solution using FRET. J Mol Biol 340:530-540. Ermolenko, D.N., Spiegel, P.C., Majumdar, Z.K., Hickerson, R.P., Clegg, R.M., and Noller, H.F. 2007. The antibiotic viomycin traps the ribosome in an intermediate state of translocation. Nat Struct Mol Biol. 6:493-7. Spiegel, P.C., Ermolenko, D.N., and Noller, H.F. 2007. Elongation factor G stabilizes the hybrid-state conformation of the 70S ribosome. RNA 13:1473-1482. Horan, L. and Noller, H.F. 2007. Intersubunit Movement is Required for Ribosomal Translocation. Proc Natl Acad Sci USA 104:4881-4885. Korostelev, A., Trakhanov, S., Laurberg, M., and Noller, H.F. 2006. Crystal Structure of a 70S Ribosome-tRNA Complex Reveals Functional Interactions and Rearrangements. Cell 126:1065-1077. Noller, H.F. 2005. RNA Structure: Reading the Ribosome. Science 309:1508-1514. Takyar, S., Hickerson, R.P. and Noller, H.F. 2005. mRNA Helicase Activity of the Ribosome. Cell 120:49-58. Noller, H. F. 2004. The driving force for molecular evolution of translation. RNA 10:1833-1837. Fredrick, K. and H. F. Noller 2003. Catalysis of ribosomal translocation by sparsomycin. Science 300:1159-62.