2BCZ

Crystal Structure of a minimal, mutant all-RNA hairpin ribozyme (U39C, G8I, 2'deoxy A-1)


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.40 Å
  • R-Value Free: 0.270 
  • R-Value Work: 0.240 
  • R-Value Observed: 0.240 

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This is version 1.3 of the entry. See complete history


Literature

Water in the Active Site of an All-RNA Hairpin Ribozyme and Effects of Gua8 Base Variants on the Geometry of Phosphoryl Transfer.

Salter, J.D.Krucinska, J.Alam, S.Grum-Tokars, V.Wedekind, J.E.

(2006) Biochemistry 45: 686-700

  • DOI: https://doi.org/10.1021/bi051887k
  • Primary Citation of Related Structures:  
    1ZFT, 1ZFV, 1ZFX, 2BCY, 2BCZ, 2FGP, 2OUE

  • PubMed Abstract: 

    The hairpin ribozyme requires functional group contributions from G8 to assist in phosphodiester bond cleavage. Previously, replacement of G8 by a series of nucleobase variants showed little effect on interdomain docking, but a 3-250-fold effect on catalysis. To identify G8 features that contribute to catalysis within the hairpin ribozyme active site, structures for five base variants were determined by X-ray crystallography in a resolution range between 2.3 and 2.7 A. For comparison, a native all-RNA "G8" hairpin ribozyme structure was refined to 2.05 A resolution. The native structure revealed a scissile bond angle (tau) of 158 degrees, which is close to the requisite 180 degrees "in-line" geometry. Mutations G8(inosine), G8(diaminopurine), G8(aminopurine), G8(adenosine), and G8(uridine) folded properly, but exhibited nonideal scissile bond geometries (tau ranging from 118 degrees to 93 degrees) that paralleled their diminished solution activities. A superposition ensemble of all structures, including a previously described hairpin ribozyme-vanadate complex, indicated the scissile bond can adopt a variety of conformations resulting from perturbation of the chemical environment and provided a rationale for how the exocyclic amine of nucleobase 8 promotes productive, in-line geometry. Changes at position 8 also caused variations in the A-1 sugar pucker. In this regard, variants A8 and U8 appeared to represent nonproductive ground states in which their 2'-OH groups mimicked the pro-R, nonbridging oxygen of the vanadate transition-state complex. Finally, the results indicated that ordered water molecules bind near the 2'-hydroxyl of A-1, lending support to the hypothesis that solvent may play an important role in the reaction.


  • Organizational Affiliation

    Department of Biochemistry and Biophysics, University of Rochester School of Medicine and Dentistry, Rochester, New York 14642, USA.


Macromolecules

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Entity ID: 1
MoleculeChains LengthOrganismImage
5'-R(*UP*CP*CP*CP*(DA)P*GP*UP*CP*CP*AP*CP*CP*G)-3')13N/A
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Entity ID: 2
MoleculeChains LengthOrganismImage
5'-R(*CP*GP*GP*UP*GP*AP*IP*AP*AP*GP*GP*G)-3'12N/A
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Entity ID: 3
MoleculeChains LengthOrganismImage
5'-R(*GP*GP*CP*AP*GP*AP*GP*AP*AP*AP*CP*AP*CP*AP*CP*GP*A)-3'17N/A
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Entity ID: 4
MoleculeChains LengthOrganismImage
5'-R(*UP*CP*GP*UP*GP*GP*UP*AP*CP*AP*UP*UP*AP*CP*CP*UP*GP*CP*C)-3'19N/A
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  • Reference Sequence
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.40 Å
  • R-Value Free: 0.270 
  • R-Value Work: 0.240 
  • R-Value Observed: 0.240 
  • Space Group: P 61 2 2
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 94.49α = 90
b = 94.49β = 90
c = 127.58γ = 120
Software Package:
Software NamePurpose
CNSrefinement
CrystalCleardata reduction
CrystalCleardata scaling
CNSphasing

Structure Validation

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Entry History 

Deposition Data

Revision History  (Full details and data files)

  • Version 1.0: 2006-02-14
    Type: Initial release
  • Version 1.1: 2008-05-01
    Changes: Version format compliance
  • Version 1.2: 2011-07-13
    Changes: Version format compliance
  • Version 1.3: 2023-08-23
    Changes: Data collection, Database references, Derived calculations, Refinement description