PDBsum entry 2d0c

Hydrolase

PDB id: 2d0c

Contents

Protein chain

308 a.a. *

Metals

_MN

Waters ×124

* Residue conservation analysis

PDB id:

2d0c

Name:

Hydrolase

Title:

Crystal structure of bst-rnase hiii in complex with mn2+

Structure:

Ribonuclease hiii. Chain: a. Synonym: rnase hiii. Engineered: yes

Source:

Geobacillus stearothermophilus. Organism_taxid: 1422. Expressed in: escherichia coli. Expression_system_taxid: 562.

Resolution:

2.60Å R-factor: 0.209 R-free: 0.274

Authors:

H.Chon,H.Matsumura,Y.Koga,K.Takano,S.Kanaya

Key ref:

H.Chon et al. (2006). Crystal structure and structure-based mutational analyses of RNase HIII from Bacillus stearothermophilus: a new type 2 RNase H with TBP-like substrate-binding domain at the N terminus. J Mol Biol, 356, 165-178. PubMed id: 16343535 DOI: 10.1016/j.jmb.2005.11.017

Date:

31-Jul-05 Release date: 18-Jul-06

Protein chain

Q6L6Q4  (Q6L6Q4_GEOSE) -  Ribonuclease HIII from Geobacillus stearothermophilus

Seq: 310 a.a.

Struc: 308 a.a.

Enzyme reactions

• Enzyme class: E.C.3.1.26.4 - ribonuclease H.
• Reaction: Endonucleolytic cleavage to 5'-phosphomonoester.

DOI no: 10.1016/j.jmb.2005.11.017

J Mol Biol 356:165-178 (2006)

PubMed id: 16343535

Crystal structure and structure-based mutational analyses of RNase HIII from Bacillus stearothermophilus: a new type 2 RNase H with TBP-like substrate-binding domain at the N terminus.

H.Chon, H.Matsumura, Y.Koga, K.Takano, S.Kanaya.

ABSTRACT

Ribonuclease HIII (Bst-RNase HIII) from the moderate thermophile Bacillus stearothermophilus is a type 2 RNase H but shows poor amino acid sequence identity with another type 2 RNase H, RNase HII. It is composed of 310 amino acid residues and acts as a monomer. Bst-RNase HIII has a large N-terminal extension with unknown function and a unique active-site motif (DEDE), both of which are characteristics common to RNases HIII. To understand the role of these N-terminal extension and active-site residues, the crystal structure of Bst-RNase HIII was determined in both metal-free and metal-bound forms at 2.1-2.6 angstroms resolutions. According to these structures, Bst-RNase HIII consists of the N-terminal domain and C-terminal RNase H domain. The structures of the N and C-terminal domains were similar to those of TATA-box binding proteins and archaeal RNases HII, respectively. The steric configurations of the four conserved active-site residues were very similar to those of other type 1 and type 2 RNases H. Single Mn and Mg ions were coordinated with Asp97, Glu98, and Asp202, which correspond to Asp10, Glu48, and Asp70 of Escherichia coli RNase HI, respectively. The mutational studies indicated that the replacement of either one of these residues with Ala resulted in a great reduction of the enzymatic activity. Overproduction, purification, and characterization of the Bst-RNase HIII derivatives with N and/or C-terminal truncations indicated that the N-terminal domain and C-terminal helix are involved in substrate binding, but the former contributes to substrate binding more greatly than the latter.

Selected figure(s)

Figure 4.
Figure 4. Stereo view of electron density around the active site of Bst-RNase HIII. The structures of (a) metal-free, ((b) and (d)) Mn2+-bound, and (c) Mg2+-bound proteins are shown. In (a), (b), and (c), the 2F[o] -F[c] map contoured at the 1.0s level is shown. In (d), the 2F[o] -F[c] map and the anomalous difference map contoured at the 4.5s and 3.5s levels are shown in blue and magenta, respectively. The red and yellow crosses indicate the water molecule and metal ion, respectively.

Figure 5.
Figure 5. The structures of the active sites of various RNases H. The side-chains of the active-site residues in the crystal structures of metal-free (green), Mn2+-bound (blue), and Mg2+-bound (magenta) Bst-RNases HIII, Tk-RNase HII (red), and E. coli RNase HI (cyan) are shown. The view direction is the same as in 1 and 3.

The above figures are reprinted by permission from Elsevier: J Mol Biol (2006, 356, 165-178) copyright 2006.

Figures were selected by an automated process.