PDBsum entry: 1sn8

Hydrolase

PDB id: 1sn8

Contents

Protein chains

87 a.a. *

80 a.a. *

Metals

_PB ×2

Waters ×229

* Residue conservation analysis

PDB id:

1sn8

Name:

Hydrolase

Title:

Crystal structure of the s1 domain of rnase e from e. Coli (pb derivative)

Structure:

Ribonuclease e. Chain: a, b. Fragment: s1 domain (residues 35-125). Synonym: rnase e. Engineered: yes

Source:

Escherichia coli. Organism_taxid: 562. Gene: rne, ams, hmp1, b1084. Expressed in: escherichia coli bl21. Expression_system_taxid: 511693.

Resolution:

2.00Å R-factor: 0.185 R-free: 0.232

Authors:

M.Schubert,R.E.Edge,P.Lario,M.A.Cook,N.C.J.Strynadka, G.A.Mackie,L.P.Mcintosh

Key ref:

M.Schubert et al. (2004). Structural characterization of the RNase E S1 domain and identification of its oligonucleotide-binding and dimerization interfaces. J Mol Biol, 341, 37-54. PubMed id: 15312761 DOI: 10.1016/j.jmb.2004.05.061

Date:

10-Mar-04 Release date: 17-Aug-04

Protein chain

P21513  (RNE_ECOLI) -  Ribonuclease E

Seq: 1061 a.a.

Struc: 87 a.a.

Protein chain

P21513  (RNE_ECOLI) -  Ribonuclease E

Seq: 1061 a.a.

Struc: 80 a.a.

Enzyme reactions

• Enzyme class: Chains A, B: E.C.3.1.26.12 - Ribonuclease E.

 Gene Ontology (GO) functional annotation 

• Biochemical function: RNA binding (1 term)

DOI no: 10.1016/j.jmb.2004.05.061

J Mol Biol 341:37-54 (2004)

PubMed id: 15312761

Structural characterization of the RNase E S1 domain and identification of its oligonucleotide-binding and dimerization interfaces.

M.Schubert, R.E.Edge, P.Lario, M.A.Cook, N.C.Strynadka, G.A.Mackie, L.P.McIntosh.

ABSTRACT

S1 domains occur in four of the major enzymes of mRNA decay in Escherichia coli: RNase E, PNPase, RNase II, and RNase G. Here, we report the structure of the S1 domain of RNase E, determined by both X-ray crystallography and NMR spectroscopy. The RNase E S1 domain adopts an OB-fold, very similar to that found with PNPase and the major cold shock proteins, in which flexible loops are appended to a well-ordered five-stranded beta-barrel core. Within the crystal lattice, the protein forms a dimer stabilized primarily by intermolecular hydrophobic packing. Consistent with this observation, light-scattering, chemical crosslinking, and NMR spectroscopic measurements confirm that the isolated RNase E S1 domain undergoes a specific monomer-dimer equilibrium in solution with a K(D) value in the millimolar range. The substitution of glycine 66 with serine dramatically destabilizes the folded structure of this domain, thereby providing an explanation for the temperature-sensitive phenotype associated with this mutation in full-length RNase E. Based on amide chemical shift perturbation mapping, the binding surface for a single-stranded DNA dodecamer (K(D)=160(+/-40)microM) was identified as a groove of positive electrostatic potential containing several exposed aromatic side-chains. This surface, which corresponds to the conserved ligand-binding cleft found in numerous OB-fold proteins, lies distal to the dimerization interface, such that two independent oligonucleotide-binding sites can exist in the dimeric form of the RNase E S1 domain. Based on these data, we propose that the S1 domain serves a dual role of dimerization to aid in the formation of the tetrameric quaternary structure of RNase E as described by Callaghan et al. in 2003 and of substrate binding to facilitate RNA hydrolysis by the adjacent catalytic domains within this multimeric enzyme.

Selected figure(s)

Figure 1.
Figure 1. Tertiary structure of RneS1. 35 --125 a, A backbone representation of the crystal structure of the Pb-derivative of RneS1 35-- 125 showing a superimposition of monomers A (yellow) and B (green) from the asymmetric unit. b, A backbone representation of the ensemble of ten NMR-derived structures of RneS1, 35-- 125 superimposed on monomer A (yellow) from the crystal structure. The secondary structure of the ensemble is highlighted with b-strands in red and helices in blue. c, A cartoon representation, using chain A, of the secondary structure of RneS1 35 -- 125 . The b-strands (arrows) are named consecutively b1 to b5, while the intervening loops are L12 through L45. As drawn, the RNA-bind- ing interface lies on the back of the molecule and the dimerization interface in front. d The amides of RneS1 35 --125 show- ing enhanced mobility on a sub-nanosecond timescale, as evident by reduced 1 H{ 15 N}-NOE values, are highlighted in color on the backbone representation of the ensemble of NMR-derived structures. Residues with 1 H{ 15 N}-NOE values of ,0.2 (highest mobility) are shown in red, between 0.2 and 0.4 in orange, between 0.4 and 0.6 in yellow, and over 0.6 in gray. Proline residues or residues where data are not available (e.g. due to spectral overlap) are identified in light blue.

Figure 7.
Figure 7. Sequence analyses of residues crucial for the OB-fold and the dimerization and oligonucleotide-binding interfaces of RneS1. 35 -- 125 (a) Alignments based on sequence (CLUSTAL W) 65 and three-dimensional structure compari- sons (DALI) 28 of a subset of cold-shock DNA-binding family OB-fold proteins, including: RNase E, RNase G and PNPase of E. coli; major cold shock proteins CspA, CspB, CspC, and CspE of E. coli; cold shock domain of the Y-Box protein YB-1 of Homo sapiens; NusA of Thermotoga maritima; archaeal RNA polymerase II RPB4/RPB7 complex of Methanococcus jannaschii; initiation translation factor IF-5a of Pyrobaculum aerophilum; ribosomal protein L2 of Haloarcula marismortui; and the ribosomal protein S17 of E. coli. Non-conserved insertions are indicated by an X. Numbers at the top correspond to amino acid residues in E. coli RNase E, and those to the right indicate the actual range for each sequence. Residues highlighted in black, dark gray and light gray are conserved in over 90%, in 60-- 90%, and in 40 -- 60% of the proteins, respectively. The secondary structure of RNase E is indicated on the top. G66 and L68 are marked

The above figures are reprinted by permission from Elsevier: J Mol Biol (2004, 341, 37-54) copyright 2004.

Figures were selected by the author.