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Graduate Student Center
for Structural Biology
Phone: (203) 432-5627
Email: schmeing@csb.yale.edu |
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Graduate Student Center
for Structural Biology
Phone: (203) 432-5627
Email: schmeing@csb.yale.edu |
Structural Studies of Ribosomes
Summary:
The ribosome is the large macromolecular machine responsible for protein synthesis. Recently, our group solved the structure of the large ribosomal subunit from Haloarcula marismortui by X-ray crystallography to atomic resolution. This structure was valuable for the insights it lent into both the mechanism of translation and complexities of RNA structure. My research has involved both these subjects, as well as the pursuit of other ribosome structures.
The K-turn: An analysis of the three-dimensional structure of the 50S ribosomal particle from Haloarcula marismortui, undertaken by myself and Dan Klein, has led to the identification of a new small RNA motif, we call the kink-turn, or K-turn. The K-turn is an asymmetric internal loop motif characterized by a kink in the phosphodiester backbone that causes a sharp turn in the RNA helix. The stems are held together by a series of hydrophobic interactions including an A-minor interaction. One of the unpaired loop nucleotides is significantly protruded, making this motif ideal for recognition by ribosomal proteins. There are six kink turns in Hma 50S, two in Tth 30S, and one each in U4 snRNA and L30e pre-mRNA. A consensus sequence we have derived allows us to predict the presence of K-turns in many RNAs, including the 5'-UTR of L10 mRNA, helix 78 of E.coli 23S RNA, and human RNase MRP.
The peptidyl transferase reaction: We use small analogues of the 3' ends of aminoacyl tRNAs to study the peptide bond formation by biochemical and structural methods. By soaking small substrates into 50S crystals, and monitoring their conversion into product, we have established that the crystalline ribosomes are catalytically active, and in a biologically relevant conformation. We have solved structures of the Haloarcula marismortui 50S complexed with analogues that represent the substrates, intermediate and products of the peptidyl transferase reaction. These structures have revealed much about how the the ribosome carries out this reaction. A theoretical model of peptide bond formation based on these structures is available in a movie form. Now also available in the more theatrical musical version in lower, and higher resolution forms. (Warning: High res is a huge file, and will take a while to download it. I'm working on making it smaller.)
The 70S translocation complex: Elongation factor G (EF-G) is the protein
factor that catalyzes the translocation of tRNAs on the ribosome during the translation cycle. I am currently trying to solve the
structure of a complex of 70S ribosomes, EF-G, tRNAs and mRNA to study the mechanism of translocation.
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A kink-turn The Haloarcula marismortui 50S Experimental electron density of a product of the peptidyl transfease reaction
Publications:
Schmeing TM, Moore PB, Steitz TA. Structures of deacylated tRNA mimics bound to the E site of the large ribosomal subunit. RNA 2003 Nov 3;9(11):1345-1352. pdf
Hansen JL, Schmeing TM, Moore PB, Steitz TA. Structural insights into peptide bond formation. Proc Natl Acad Sci U S A. 2002 Sep 3;99(18):11670-5. pdf
Schmeing TM, Seila AC, Hansen JL, Freeborn B, Soukup JK, Scaringe SA, Strobel SA, Moore PB, Steitz TA. A pre-translocational intermediate in protein synthesis observed in crystals of enzymatically active 50S subunits. Nat Struct Biol. 2002 Mar;9(3):225-30. pdf
Hansen JL, Schmeing TM, Klein DJ, Ippolito JA, Nissen P, Ban N, Moore PB, Steitz TA (2001) "Progress towards an understanding of the structure and enzymatic mechanism of the large ribosomal subunit." Cold Spring Harbor Symposia on Quantitative Biology, Volume LXVI.
Klein DJ, Schmeing TM, Moore PB, Steitz TA. The kink-turn: a new RNA secondary structure motif. EMBO J. 2001 Aug 1;20(15):4214-21. (Klein and Schmeing contributed equally to this publication) pdf
