scholarly journals Biophysical and biochemical investigations of RNA catalysis in the hammerhead ribozyme

1999 ◽  
Vol 32 (3) ◽  
pp. 241-284 ◽  
Author(s):  
William G. Scott

1. How do ribozymes work? 2412. The hammerhead RNA as a prototype ribozyme 2422.1 RNA enzymes 2422.2 Satellite self-cleaving RNAs 2422.3 Hammerhead RNAs and hammerhead ribozymes 2443. The chemical mechanism of hammerhead RNA self-cleavage 2463.1 Phosphodiester isomerization via an SN2(P) reaction 2473.2 The canonical role of divalent metal ions in the hammerhead ribozyme reaction 2513.3 The hammerhead ribozyme does not actually require metal ions for catalysis 2543.4 Hammerhead RNA enzyme kinetics 2574. Sequence requirements for hammerhead RNA self-cleavage 2604.1 The conserved core, mutagenesis and functional group modifications 2604.2 Ground-state vs. transition-state effects 2615. The three-dimensional structure of the hammerhead ribozyme 2625.1 Enzyme–inhibitor complexes 2625.2 Enzyme–substrate complex in the initial state 2645.3 Hammerhead ribozyme self-cleavage in the crystal 2645.4 The requirement for a conformational change 2655.5 Capture of conformational intermediates using crystallographic freeze-trapping 2665.6 The structure of a hammerhead ribozyme ‘early’ conformational intermediate 2675.7 The structure of a hammerhead ribozyme ‘later’ conformational intermediate 2685.8 Is the conformational change pH dependent? 2695.9 Isolating the structure of the cleavage product 2715.10 Evidence for and against additional large-scale conformation changes 2745.11 NMR spectroscopic studies of the hammerhead ribozyme 2786. Concluding remarks 2807. Acknowledgements 2818. References 2811. How do ribozymes work? 241The discovery that RNA can be an enzyme (Guerrier-Takada et al. 1983; Zaug & Cech, 1986) has created the fundamental question of how RNA enzymes work. Before this discovery, it was generally assumed that proteins were the only biopolymers that had sufficient complexity and chemical heterogeneity to catalyze biochemical reactions. Clearly, RNA can adopt sufficiently complex tertiary structures that make catalysis possible. How does the three- dimensional structure of an RNA endow it with catalytic activity? What structural and functional principles are unique to RNA enzymes (or ribozymes), and what principles are so fundamental that they are shared with protein enzymes?

2009 ◽  
Vol 284 (24) ◽  
pp. 16126-16134 ◽  
Author(s):  
Sarah L. Greig ◽  
Mazdak Radjainia ◽  
Alok K. Mitra

Colicin Ia is a soluble, harpoon-shaped bacteriocin which translocates across the periplasmic space of sensitive Escherichia coli cell by parasitizing an outer membrane receptor and forms voltage-gated ion channels in the inner membrane. This process leads to cell death, which has been thought to be caused by a single colicin Ia molecule. To directly visualize the three-dimensional structure of the channel, we generated two-dimensional crystals of colicin Ia inserted in lipid-bilayer membranes and determined a ∼17 three-dimensional model by electron crystallography. Supported by velocity sedimentation, chemical cross-linking and single-particle image analysis, the three-dimensional structure is a crown-shaped oligomer enclosing a ∼35 Å-wide extrabilayer vestibule. Our study suggests that lipid insertion instigates a global conformational change in colicin Ia and that more than one molecule participates in the channel architecture with the vestibule, possibly facilitating the known large scale peptide translocation upon channel opening.


We attempt to catalogue those features of the three-dimensional structure of the Earth that are well-constrained by low-frequency data (i.e. periods longer than about 125 seconds). The dominant signals in such data are the surface-wave equivalent modes whose phase characteristics are mainly affected by a large scale structure of harmonic degree two in the upper mantle. Available aspherical models predict this phase behaviour quite well, but do not give an accurate prediction of the observed waveforms and we must appeal to higher-order structure an d /o r coupling effects to give the observed complexity of the data. Strong splitting of modes which sample the cores of the Earth is also observed and, though we do not yet have a model of aspherical structure which gives quantitative agreement with these data, anisotropy or large-scale aspherical structure in the inner core appears to be required to model the observed signal.


Author(s):  
Hironori UENO ◽  
Khanh Huy BUI ◽  
Takuji ISHIKAWA ◽  
Takami YAMAGUCHI ◽  
Takashi ISHIKAWA

Several different kinds of seismological data, spanning more than three orders of magnitude in frequency, have been employed in the study of the Earth’s large-scale three-dimensional structure. These yield different but overlapping information, which is leading to a coherent picture of the Earth’s internal heterogeneity. In this article we describe several methods of seismic inversion and intercom pare the resulting models. Models of upper-mantle shear velocity based upon mantle waveforms (Woodhouse & Dziewonski ( J. geophys. Res . 89 , 5953-5986 (1984))) ( f ≲ 7 mHz) and long-period body waveforms ( f ≲ 20 mHz; Woodhouse & Dziewonski ( Eos, Wash . 67 , 307 (1986))) show the mid-oceanic ridges to be the major low-velocity anomalies in the uppermost mantle, together with regions in the western Pacific, characterized by back-arc volcanism. High velocities are associated with the continents, and in particular with the continental shields, extending to depths in excess of 300 km. By assuming a given ratio between density and wave velocity variations, and a given mantle viscosity structure, such models have been successful in explaining some aspects of observed plate motion in terms of thermal convection in the mantle (Forte & Peltier ( J. geophys. Res . 92 , 3645-3679 (1987))). An im portant qualitative conclusion from such analysis is that the magnitude of the observed seismic anomalies is of the order expected in a convecting system having the viscosity, tem perature derivatives and flow rates which characterize the mantle. Models of the lower mantle based upon P-wave arrival times ( f ≈ 1 Hz; Dziewonski ( J. geophys. Res . 89 , 5929-5952 (1984)); Morelli & Dziewonski ( Eos, Wash . 67 , 311 (1986))) SH waveforms ( f ≈ mHz; Woodhouse & Dziewonski (1986)) and free oscillations (Giardini et al . ( Nature, Lond . 325 , 405-411 (1987); J. geophys. Res. 93 , 13716—13742 (1988))) ( f ≈ 0.5-5 mHz) show a very long wavelength pattern, largely contained in spherical harmonics of degree 2, which is present over a large range of depths (1000-2700 km). This anomaly has been detected in both compressional and shear wave velocities, and yields a ratio of relative perturbations in v s and v P in the lower mantle in the range 2-2.5. Such values, which are much larger than has sometimes been assumed, roughly correspond to the case that perturbations in shear modulus dominate those in bulk modulus. It is this anomaly that is mainly responsible for the observed low-degree geoid undulations (Hager et al. Nature, Lond . 313 , 541-545 (1985))). In the upper part of the lower mantle this pattern consists of a high-velocity feature skirting the subduction zones of the Pacific and extending from Indonesia to the Mediterranean, with low velocities elsewhere; thus it appears to be associated with plate convergence and subduction. The pattern of wave speeds in the lowermost mantle is such that approximately 80% of hot spots are in regions of lower than average velocities in the D" region. The topography of the core-mantle boundary, determined from the arrival times of reflected and transmitted waves (Morelli & Dziewonski ( Nature, Lond . 325 , 678-683 (1987))), exhibits a pattern of depressions encircling the Pacific, having an amplitude of approximately ± 5 km, which has been shown to be consistent with the stresses induced by density anomalies inferred from tom ographic models of the lower mantle (Forte & Peltier ( Tectonphysics (In the press.) (1989))). By using both free oscillations (Woodhouse et al . ( Geophys. Res. Lett . 13 , 1549-1552 (1986))) and travel-time data (Morelli et al . ( Geophys. Res. Lett . 13 , 1545—1548 (1986))), the inner core has been found to be anisotropic, exhibiting high velocities for waves propagating parallel to the Earth ’s rotation axis and low velocities in the equatorial plane. Tomographic models represent an instantaneous, low-resolution image of a convecting system. They require for their detailed interpretation knowledge of mineral and rock properties that are, as yet, poorly known but that laboratory experiments can potentially determ ine. The fact that the present distribution of seismic anomalies must represent the current configuration of therm al and compositional heterogeneity advected by m antle flow, imposes a complex set of constraints on the possible modes of convection in the m antle of which the implications have not yet been worked out; this will require num erical modelling of convection in three dimensions, which only recently has become feasible. Thus the interpretation of the ‘geographical’ information from seismology in terms of geodynamical processes is a matter of considerable complexity, and we may expect that a number of the conclusions to be drawn from the seismological results lie in the future.


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