Ribosome crystallography

Yonath, A. (2005) Ribosomal Crystallography Peptide Bond Formation, Chaperone Assistance and Antibiotics Activity. Mol. Cells, 20, 1-16. 

Abstract Ribosome crystallography has recently been the subject of the Nobel Prize in... 

Virus and ribosome crystallography is one of the main uses of SR and one which takes advantage of a combination of SR properties such as the high flux, the fine collimation and short wavelengths. 

The combination of all the advantages of SR is especially needed in virus and ribosome crystallography where the unit cells are very large. Data collection from very large unit cell constant crystals benefits from... 

This is a very important project in structural molecular biology. As was stated at the beginning of this section, ribosome crystallography has been made possible by SR, used in conjunction with freezing of the crystals to preserve their lifetime in the beam at high resolution. 

The synchrotron should not be seen as a replacement for facilities in the home laboratory but as a means for meeting technically challenging data collection problems. Of course, in the absence of any home X-ray facilities, the central facility can be used but this is not terribly efficient because of the necessity of long-term scheduling of many users. Hence, characterisation of samples (e.g. of heavy atom derivatives) should be done at home unless the project is entirely reliant on the SR source in this category are many virus studies and ribosome crystallography as well as small crystal projects. 

The large unit cell cases, i.e. mammalian viruses and ribosome crystallography, have slotted in extremely well at SR centres because of the intrinsic weakness of scattering of these samples and the great difficulties in working with them in the home laboratory. 

The structure of the ribosome s large subunit has recently been resolved at 2.4-A resolution by means of x-ray crystallography N Ban, P Nissen, J Hansen, PB Moore, TA Steitz. Science... 

Aminoglycosides remain clinically important antibiotics. NMR provided the initial breakthrough in structural understanding of aminoglycoside action on the ribosome, and it remains a powerful tool for the biophysical characterization of drug-RNA interaction. The combined use of NMR, X-ray crystallography, thermodynamic and functional assays, and computational methods is needed to drive forward the development of new aminoglycosides with improved clinical properties. The rich data described above, combined with the application of new synthetic methods, bode well for the future. 

The sequences of all three pieces of RNA in the E. coli ribosomes are known as are those from many other species. These include eukaryotic mitochondrial, plas-tid, and cytosolic rRNA. From the sequences alone, it was clear that these long molecules could fold into a complex series of hairpin loops resembling those in tRNA. For example, the 16S rRNA of E. coli can fold as in Fig. 29-2A and eukaryotic 18S RNA in a similar way (Fig. 29-4).38/39/67 69 The actual secondary structures of 16S and 18S RNAs, within the folded molecules revealed by X-ray crystallography, are very similar to that shown in Fig. 29-2A. Ribosomal RNAs undergo many posttranscriptional alterations. Methylation of 2 -hydroxyls and of the nucleic acid bases as well as conversion to pseudouridines (pp. 1638-1641) predominate over 200 modifications, principally in functionally important locations that have been found in human rRNA.69a... 

This chapter provides a simplified overview of how researchers use the technique of X-ray crystallography to learn macromolecular structures. Chapters 3-8 are simply expansions of the material in this chapter. In order to keep the language simple, I will speak primarily of proteins, but the concepts I describe apply to all macromolecules and macromolecular assemblies that possess ordered structure, including carbohydrates, nucleic acids, and nucleo-protein complexes like ribosomes and whole viruses. 

In principle the global structure of an RNA junction could be determined when it is complexed with bound protein. This has not been accomplished to date in RNA, but it has proven itself for DNA junctions. For example, the structure of DNA junctions bound by the junctionresolving enzymes T4 endonuclease VII (Pohler et al., 1996) and T7 endonuclease I (Declais et al., 2003) have both been determined by comparative gel electrophoresis. It was found that both proteins substantially alter the global shape of the DNA junction in different ways. These structures were both recendy confirmed by X-ray crystallography (Biertiimpfel et al., 2007 Hadden et al., 2007), showing that comparative gel electrophoresis functions reliably for protein complexes. There is no reason to expect that the method would not work equally well for RNA junction complexes. The binding of the ribosomal protein SI 5 to a three-way RNA junction has been studied by an electrophoretic approach that is related to comparative gel electrophoresis (Batey and Williamson, 1998). 

The halobhilic proteins and macromolecular complexes on which structural studies are in progress include ferredoxin from H. maris-mortui (X-ray crystallography), ribosomal subunits from H. marismor-... 

MeSH or H2S give C Hg(SMe) 4 and CHg4S2 , respectively.130 Several of these compounds have been used as sources of heavy-atoms in X-ray crystallography of macromolecules such as nucleic acids, membrane proteins and ribosomes.131 133... 

Like the aminoglycosides, the binding site of the macrolide antibiotics with the large ribosomal subunit has also been determined to atomic resolution by X-ray crystallography (29, 38, 39). Key interactions between the antibiotic and the 23 S rRNA occur and are mediated through the essential desosamine sugar ... 

Figure 29.17. Ribosomal RNA Folding Pattern. (A) The secondary structure of 16S ribosomal RNA deduced from sequence comparison and the results of chemical studies. (B) The tertiary structure of 16S RNA determined hy x-ray crystallography. [Part A courtesy of Dr. Bryn Weiser and Dr. Harry Noller.]...

Michael Rossmann and Ada Yonat. Rossmann did important work on virus structures and made major contributions in crystallography over a long period of time. Ada Yonat for ribosome and for her work over a period of many years, her perseverance. It reminds me somewhat of my own experience. 

Keywords ribosome antibiotics RNA drug design molecular modeling crystallography docking QSAR Macrolides Oxazolidinones aminoglycosides molecular properties... 

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