James M Ogle

University of Oxford, Oxford, England, United Kingdom

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Publications (7)117.2 Total impact

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    James M Ogle · V Ramakrishnan
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    ABSTRACT: The underlying basis for the accuracy of protein synthesis has been the subject of over four decades of investigation. Recent biochemical and structural data make it possible to understand at least in outline the structural basis for tRNA selection, in which codon recognition by cognate tRNA results in the hydrolysis of GTP by EF-Tu over 75 A away. The ribosome recognizes the geometry of codon-anticodon base pairing at the first two positions but monitors the third, or wobble position, less stringently. Part of the additional binding energy of cognate tRNA is used to induce conformational changes in the ribosome that stabilize a transition state for GTP hydrolysis by EF-Tu and subsequently result in accelerated accommodation of tRNA into the peptidyl transferase center. The transition state for GTP hydrolysis is characterized, among other things, by a distorted tRNA. This picture explains a large body of data on the effect of antibiotics and mutations on translational fidelity. However, many fundamental questions remain, such as the mechanism of activation of GTP hydrolysis by EF-Tu, and the relationship between decoding and frameshifting.
    Full-text · Article · Feb 2005 · Annual Review of Biochemistry
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    James M Ogle · Andrew P Carter · V Ramakrishnan
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    ABSTRACT: During the decoding process, tRNA selection by the ribosome is far more accurate than expected from codon-anticodon pairing. Antibiotics such as streptomycin and paromomycin have long been known to increase the error rate of translation, and many mutations that increase or lower accuracy have been characterized. Recent crystal structures show that the specific recognition of base-pairing geometry leads to a closure of the domains of the small subunit around cognate tRNA. This domain closure is likely to trigger subsequent steps in tRNA selection. Many antibiotics and mutations act by making the domain closure more or less favourable. In conjunction with recent cryoelectron microscopy structures of the ribosome, a comprehensive structural understanding of the decoding process is beginning to emerge.
    Full-text · Article · Jun 2003 · Trends in Biochemical Sciences
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    ABSTRACT: A structural and mechanistic explanation for the selection of tRNAs by the ribosome has been elusive. Here, we report crystal structures of the 30S ribosomal subunit with codon and near-cognate tRNA anticodon stem loops bound at the decoding center and compare affinities of equivalent complexes in solution. In ribosomal interactions with near-cognate tRNA, deviation from Watson-Crick geometry results in uncompensated desolvation of hydrogen-bonding partners at the codon-anticodon minor groove. As a result, the transition to a closed form of the 30S induced by cognate tRNA is unfavorable for near-cognate tRNA unless paromomycin induces part of the rearrangement. We conclude that stabilization of a closed 30S conformation is required for tRNA selection, and thereby structurally rationalize much previous data on translational fidelity.
    Full-text · Article · Dec 2002 · Cell
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    Preview · Article · Feb 2002 · Biochemical Society Transactions
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    ABSTRACT: Isopenicillin N synthase (IPNS) catalyses formation of bicyclic isopenicillin N, precursor to all penicillin and cephalosporin antibiotics, from the linear tripeptide delta-(L-alpha-aminoadipoyl)-L-cysteinyl-D-valine. IPNS is a non-haem iron(II)-dependent enzyme which utilises the full oxidising potential of molecular oxygen in catalysing the bicyclisation reaction. The reaction mechanism is believed to involve initial formation of the beta-lactam ring (via a thioaldehyde intermediate) to give an iron(IV)-oxo species, which then mediates closure of the 5-membered thiazolidine ring. Here we report experiments employing time-resolved crystallography to observe turnover of an isosteric substrate analogue designed to intercept the catalytic pathway at an early stage. Reaction in the crystalline enzyme-substrate complex was initiated by the application of high-pressure oxygen, and subsequent flash freezing allowed an oxygenated product to be trapped, bound at the iron centre. A mechanism for formation of the observed thiocarboxylate product is proposed. In the absence of its natural reaction partner (the N-H proton of the L-cysteinyl-D-valine amide bond), the proposed hydroperoxide intermediate appears to attack the putative thioaldehyde species directly. These results shed light on the events preceding beta-lactam closure in the IPNS reaction cycle, and enhance our understanding of the mechanism for reaction of the enzyme with its natural substrate.
    Full-text · Article · Jan 2002 · Chemistry & Biology
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    ABSTRACT: Crystal structures of the 30S ribosomal subunit in complex with messenger RNA and cognate transfer RNA in the A site, both in the presence and absence of the antibiotic paromomycin, have been solved at between 3.1 and 3.3 angstroms resolution. Cognate transfer RNA (tRNA) binding induces global domain movements of the 30S subunit and changes in the conformation of the universally conserved and essential bases A1492, A1493, and G530 of 16S RNA. These bases interact intimately with the minor groove of the first two base pairs between the codon and anticodon, thus sensing Watson-Crick base-pairing geometry and discriminating against near-cognate tRNA. The third, or "wobble," position of the codon is free to accommodate certain noncanonical base pairs. By partially inducing these structural changes, paromomycin facilitates binding of near-cognate tRNAs.
    Full-text · Article · Jun 2001 · Science

  • No preview · Article · Feb 2001 · Cold Spring Harbor Symposia on Quantitative Biology