PDBsum entry 1zd7

Transferase

PDB id: 1zd7

Contents

Protein chains

159 a.a. *

Ligands

SO4 ×4

Waters ×240

* Residue conservation analysis

PDB id:

1zd7

Name:

Transferase

Title:

1.7 angstrom crystal structure of post-splicing form of a dnae intein from synechocystis sp. Pcc 6803

Structure:

DNA polymerase iii alpha subunit. Chain: a, b. Fragment: residues 1-159. Synonym: dnae intein. Engineered: yes

Source:

Synechocystis sp.. Organism_taxid: 1148. Strain: pcc 6803. Expressed in: escherichia coli. Expression_system_taxid: 562.

Resolution:

1.70Å R-factor: 0.177 R-free: 0.202

Authors:

P.Sun,S.Ye,S.Ferrandon,T.C.Evans,M.Q.Xu,Z.Rao

Key ref:

P.Sun et al. (2005). Crystal structures of an intein from the split dnaE gene of Synechocystis sp. PCC6803 reveal the catalytic model without the penultimate histidine and the mechanism of zinc ion inhibition of protein splicing. J Mol Biol, 353, 1093-1105. PubMed id: 16219320 DOI: 10.1016/j.jmb.2005.09.039

Date:

14-Apr-05 Release date: 24-Jan-06

Protein chains

P74750  (DPO3A_SYNY3) -  DNA polymerase III subunit alpha (Fragment) from Synechocystis sp. (strain ATCC 27184 / PCC 6803 / Kazusa)

Seq: 1355 a.a.

Struc: 159 a.a.

Enzyme reactions

• Enzyme class: E.C.2.7.7.7 - DNA-directed Dna polymerase.
• Reaction: DNA(n) + a 2'-deoxyribonucleoside 5'-triphosphate = DNA(n+1) + diphosphate

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

Added reference

DOI no: 10.1016/j.jmb.2005.09.039

J Mol Biol 353:1093-1105 (2005)

PubMed id: 16219320

Crystal structures of an intein from the split dnaE gene of Synechocystis sp. PCC6803 reveal the catalytic model without the penultimate histidine and the mechanism of zinc ion inhibition of protein splicing.

P.Sun, S.Ye, S.Ferrandon, T.C.Evans, M.Q.Xu, Z.Rao.

ABSTRACT

The first naturally occurring split intein was found in the dnaE gene of Synechocystis sp. PCC6803 and belongs to a subclass of inteins without a penultimate histidine residue. We describe two high-resolution crystal structures, one derived from an excised Ssp DnaE intein and the second from a splicing-deficient precursor protein. The X-ray structures indicate that His147 in the conserved block F activates the side-chain N(delta) atom of the intein C-terminal Asn159, leading to a nucleophilic attack on the peptide bond carbonyl carbon atom at the C-terminal splice site. In this process, Arg73 appears to stabilize the transition state by interacting with the carbonyl oxygen atom of the scissile bond. Arg73 also seems to substitute for the conserved penultimate histidine residue in the formation of an oxyanion hole, as previously identified in other inteins. The finding that the precursor structure contains a zinc ion chelating the highly conserved Cys160 and Asp140 reveals the structural basis of Zn2+-mediated inhibition of protein splicing. Furthermore, it is of interest to observe that the carbonyl carbon atom of Asn159 and N(eta) of Arg73 are 2.6 angstroms apart in the free intein structure and 10.6 angstroms apart in the precursor structure. The orientation change of the aromatic ring of Tyr-1 following the initial acyl shift may be a key switching event contributing to the alignment of Arg73 and the C-terminal scissile bond, and may explain the sequential reaction property of the Ssp DnaE intein.

Selected figure(s)

Figure 1.
Figure 1. (a) Diagram of conserved intein motifs of bifunctional inteins, mini inteins and the Ssp DnaE split intein. Blocks A and B (black) in the N-terminal subdomain (magenta) and blocks F and G (black) in the C-terminal subdomain (yellow) are shared by the splicing domains and the endonuclease domain is shown in grey. Residues involved in nucleophilic attack (letters in a box), as well as other highly conserved amino acids are indicated below the block diagram. (b) A representation of the Ssp DnaE intein fusions. The exDnaE fusion protein consists of maltose-binding protein (MBP), the full-length wild-type Ssp DnaE intein (residues 1-159, which include 123 amino acid residues from the N terminus and 36 amino acid residues from the C terminus) with five native extein residues at its N terminus and three native residues at its C terminus, and the CBD. The resulting protein exDnaE is splicing functional. Black arrows indicate the splicing sites of Ssp DnaE intein. preDnaE consists of CBD, Ssp DnaB intein and the full-length Ssp DnaE intein with C1A and N159A mutations (residue 1-159) along with five native extein residues at its N terminus and three native residues at its C terminus. The black arrow shows the cleavage site of the modified Ssp DnaB intein.41 The intein proteins after purification are indicated in red.

Figure 3.
Figure 3. (a) Close-up stereo view of the superposition of the N-terminal subdomain of preDnaE (cyan) and exDnaE (purple). (b) Stereo view of the modeled N-terminal catalytic module of Ssp DnaE intein. (e) Close-up stereo view of the superposition of the C-terminal subdomain of preDnaE (cyan) and exDnaE (purple). (f) Stereo view of the modeled C-terminal catalytic module of Ssp DnaE intein. Residues are shown in ball-and-stick representations. The broken lines indicate hydrogen bonds, and bond distances are labeled. (c), (d), (g), (h) and (i) A chemical mechanism proposed for splicing the Ssp DnaE intein. The red arrows indicate the routes of nucleophilic attacks in the splicing pathway. Broken lines indicate hydrogen bonds. The tetrahedral intermediate formed by an N-S acyl rearrangement at Cys1 is not shown.

The above figures are reprinted by permission from Elsevier: J Mol Biol (2005, 353, 1093-1105) copyright 2005.

Figures were selected by an automated process.