PDBsum entry 2drb

Transferase/RNA

PDB id: 2drb

Contents

Protein chain

437 a.a. *

DNA/RNA

Ligands

SO4 ×2

Waters ×112

* Residue conservation analysis

PDB id:

2drb

Name:

Transferase/RNA

Title:

Complex structure of cca-adding enzyme with trnaminicca

Structure:

tRNA (35-mer). Chain: b. Engineered: yes. Cca-adding enzyme. Chain: a. Synonym: tRNA nucleotidyltransferase, tRNA adenylyl-/cytidylyl- transferase, tRNA cca-pyrophosphorylase, tRNA-nt. Engineered: yes

Source:

Synthetic: yes. Archaeoglobus fulgidus. Organism_taxid: 2234. Expressed in: escherichia coli. Expression_system_taxid: 562

Biol. unit:

Tetramer (from PQS)

Resolution:

2.80Å R-factor: 0.215 R-free: 0.282

Authors:

K.Tomita,R.Ishitani,S.Fukai,O.Nureki

Key ref:

K.Tomita et al. (2006). Complete crystallographic analysis of the dynamics of CCA sequence addition. Nature, 443, 956-960. PubMed id: 17051158 DOI: 10.1038/nature05204

Date:

08-Jun-06 Release date: 14-Nov-06

Protein chain

O28126  (CCA_ARCFU) -  CCA-adding enzyme from Archaeoglobus fulgidus (strain ATCC 49558 / DSM 4304 / JCM 9628 / NBRC 100126 / VC-16)

Seq: 437 a.a.

Struc: 437 a.a.

DNA/RNA chain

G-G-C-C-C-G-G-G-G-C-G-G-U-U-C-G-A-U-U-C-C-G-C-C-C-U-G-G-G-C-C-A-C-C-A 35 bases

Enzyme reactions

• Enzyme class: E.C.2.7.7.72 - Cca tRNA nucleotidyltransferase.
• Reaction: a tRNA precursor + 2 CTP + ATP = a tRNA with a 3' CCA end + 3 diphosphate

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

reference

DOI no: 10.1038/nature05204

Nature 443:956-960 (2006)

PubMed id: 17051158

Complete crystallographic analysis of the dynamics of CCA sequence addition.

K.Tomita, R.Ishitani, S.Fukai, O.Nureki.

ABSTRACT

CCA-adding polymerase matures the essential 3'-CCA terminus of transfer RNA without any nucleic-acid template. However, it remains unclear how the correct nucleotide triphosphate is selected in each reaction step and how the polymerization is driven by the protein and RNA dynamics. Here we present complete sequential snapshots of six complex structures of CCA-adding enzyme and four distinct RNA substrates with and without CTP (cytosine triphosphate) or ATP (adenosine triphosphate). The CCA-lacking RNA stem extends by one base pair to force the discriminator nucleoside into the active-site pocket, and then tracks back after incorporation of the first cytosine monophosphate (CMP). Accommodation of the second CTP clamps the catalytic cleft, inducing a reorientation of the turn, which flips C74 to allow CMP to be accepted. In contrast, after the second CMP is added, the polymerase and RNA primer are locked in the closed state, which directs the subsequent A addition. Between the CTP- and ATP-binding stages, the side-chain conformation of Arg 224 changes markedly; this is controlled by the global motion of the enzyme and position of the primer terminus, and is likely to achieve the CTP/ATP discrimination, depending on the polymerization stage. Throughout the CCA-adding reaction, the enzyme tail domain firmly anchors the TPsiC-loop of the tRNA, which ensures accurate polymerization and termination.

Selected figure(s)

Figure 2.
Figure 2: Expansion and contraction of the primer RNA helix at the mini-D stage. a, Extended mini-helix structure at the mini-D stage. The simulated annealed omit maps (contoured at 3.5 ) for the indicated nucleosides are shown. b, Back-tracked mini-helix structure at the mini-DC. Other stages have the same standard helix structure.

Figure 3.
Figure 3: Active-site structure at each reaction stage. a, Mini-D stage (the simulated annealed omit maps contoured at 4 for G1 and A73 are shown). b, Mini-DC stage. c, Mini-DC + CTP stage. d, Mini-DCC stage. e, Mini-DCC + ATP stage. f, Mini-DCCA stage. g, Mature tRNA dissociation stage (PDB ID: 1SZ1)^14. The acceptor stem expansion at the first CCA-adding step can easily be seen by the fact that Asp 291 recognizes the 2'-OH group of C72 in the mini-D stage, but A73 in the other stages. The catalytic triad comprises Glu 59, Asp 61 and Asp 110. The hydrogen bonds are represented by dotted lines.

The above figures are reprinted by permission from Macmillan Publishers Ltd: Nature (2006, 443, 956-960) copyright 2006.

Figures were selected by an automated process.