PDBsum entry 1vj7

Hydrolase, transferase

PDB id: 1vj7

Contents

Protein chains

326 a.a. *

310 a.a. *

Ligands

GDP ×2

GPX

Metals

_MN ×2

Waters ×196

* Residue conservation analysis

PDB id:

1vj7

Name:

Hydrolase, transferase

Title:

Crystal structure of the bifunctional catalytic fragment of relseq, the rela/spot homolog from streptococcus equisimilis.

Structure:

Bifunctional rela/spot. Chain: a, b. Fragment: (p)ppgpp-3'-pyrophosphohydrolase and (p)ppgpp-synthetase subdomains. Synonym: atp:gtp 3'- pyrophosphotransferase, ppgpp synthetase i, p, ppgpp synthetase, stringent response-like protein. Engineered: yes

Source:

Streptococcus dysgalactiae subsp. Equisimilis. Organism_taxid: 119602. Strain: subsp. Equisimilis. Gene: rela, rel, spot, rsh. Expressed in: escherichia coli. Expression_system_taxid: 562.

Resolution:

2.10Å R-factor: 0.238 R-free: 0.272

Authors:

T.Hogg,U.Mechold,H.Malke,M.Cashel,R.Hilgenfeld

Key ref:

T.Hogg et al. (2004). Conformational antagonism between opposing active sites in a bifunctional RelA/SpoT homolog modulates (p)ppGpp metabolism during the stringent response [corrected]. Cell, 117, 57-68. PubMed id: 15066282 DOI: 10.1016/S0092-8674(04)00260-0

Date:

03-Feb-04 Release date: 04-May-04

Protein chain

Q54089  (RELA_STREQ) -  Bifunctional (p)ppGpp synthase/hydrolase RelA from Streptococcus dysgalactiae subsp. equisimilis

Seq: 739 a.a.

Struc: 326 a.a.*

Protein chain

Q54089  (RELA_STREQ) -  Bifunctional (p)ppGpp synthase/hydrolase RelA from Streptococcus dysgalactiae subsp. equisimilis

Seq: 739 a.a.

Struc: 310 a.a.*

* PDB and UniProt seqs differ at 14 residue positions (black crosses)

Enzyme reactions

• Enzyme class 2: Chains A, B: E.C.2.7.6.5 - Gtp diphosphokinase.
• Reaction: GTP + ATP = guanosine 3'-diphosphate 5'-triphosphate + AMP

GTP + ATP (Bound ligand (Het Group name = GDP) matches with 87.50% similarity) = guanosine 3'-diphosphate 5'-triphosphate + AMP (Bound ligand (Het Group name = GPX) matches with 77.50% similarity)

• Enzyme class 3: Chains A, B: E.C.3.1.7.2 - guanosine-3',5'-bis(diphosphate) 3'-diphosphatase.
• Reaction: guanosine 3',5'-bis(diphosphate) + H2O = GDP + diphosphate + H⁺

guanosine 3',5'-bis(diphosphate) (Bound ligand (Het Group name = GPX) matches with 86.11% similarity) + H2O = GDP + diphosphate (Bound ligand (Het Group name = GDP) corresponds exactly) + H(+)

Note, where more than one E.C. class is given (as above), each may correspond to a different protein domain or, in the case of polyprotein precursors, to a different mature protein.

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

reference

DOI no: 10.1016/S0092-8674(04)00260-0

Cell 117:57-68 (2004)

PubMed id: 15066282

Conformational antagonism between opposing active sites in a bifunctional RelA/SpoT homolog modulates (p)ppGpp metabolism during the stringent response [corrected].

T.Hogg, U.Mechold, H.Malke, M.Cashel, R.Hilgenfeld.

ABSTRACT

Enzymes of the Rel/Spo family enable bacteria to survive prolonged periods of nutrient limitation by producing an intracellular signaling alarmone, (p)ppGpp, which triggers the so-called stringent response. Both the synthesis of (p)ppGpp from ATP and GDP(GTP), and its hydrolysis to GDP(GTP) and pyrophosphate, are catalyzed by Rel/Spo proteins. The 2.1 A crystal structure of the bifunctional catalytic fragment of the Rel/Spo homolog from Streptococcus dysgalactiae subsp. equisimilis, Rel(Seq), reveals two conformations of the enzyme corresponding to known reciprocal activity states: (p)ppGpp-hydrolase-OFF/(p)ppGpp-synthetase-ON and hydrolase-ON/synthetase-OFF. The hydrolase and synthetase domains bear remarkable similarities to the catalytic domains of the cyclic phosphodiesterase and nucleotidyltransferase superfamilies, respectively. The active sites, separated by more than 30 A, contain bound nucleotides including an unusual (p)ppGpp derivative, GDP-2':3'-cyclic monophosphate. Reciprocal regulation of the antagonistic catalytic activities, suggested by the structure, is supported by mutagenesis experiments and appears to involve ligand-induced signal transmission between the two active sites.

Selected figure(s)

Figure 2.
Figure 2. Similarities between the Catalytic Domains of Rel[Seq], Human Phosphodiesterase (PDE) and Human DNA Polymerase Beta (pol β)Structural and topological diagrams highlighting equivalent folds and active-site residues for: (A) catalytic domain (residues 152−528) of PDE4; (B) Rel[Seq]1–385; (C) catalytic domain (residues 10−335) of pol β. Homologous structural elements are displayed as ribbons; nonequivalent regions as thin gray lines. Monomer 2 of Rel[Seq]1–385 is shown, with ppG2′:3′p, and GDP. Dark blue sphere, catalytic metal ion (Zn^2+ for PDE4; Mn^2+ for Rel[Seq]). Conserved residues of the H−X[(n)]−HD−X[(n)]−D metal binding tetrad are labeled in the accompanying topology diagrams (A and B). Two of the three catalytic carboxylates in pol β (Asp190 and Asp256, C) are also found in Rel[Seq] (Asp264 and Glu323, B). Rel/Spo enzymes lack a counterpart for the second carboxylate of the D-X-D motif in NTases (Asp192 in pol β).

Figure 4.
Figure 4. Conformations of the Synthetase Site in Rel[Seq]1–385 and a Superposition with pol β(A) The synthetase-ON conformation (monomer 1). Coloring is according to Figure 1; individual structural elements are labeled in red. The nucleophilic O3′ of GDP is marked. The final 2mFo-DFc electron density map, shown for GDP (blue mesh), is contoured at 1.0 σ. Selected H-bonds are shown as gray dashed lines. The catalytic loop (α13/β4) harboring Asp264 is stabilized in a 3[10]-helical conformation through multiple van der Waals interactions (represented by black double-arrow dashed lines) with loop α11/α12 and the first two turns of α12 (labeled t1, t2). Chain traces extending from loops α11/α12, β3/α13, and α13/β4 are not visible due to image slab restrictions.(B) The synthetase-OFF conformation (monomer 2). The α11/α12 loop and the first two turns of α12 are disordered; the resulting elimination of van der Waals contacts to the catalytic loop (α13/β4) leads to (1), partial refolding of the latter into an N-terminal extension of β4, and (2), disorder of residues 254–261 including the remaining residues of the catalytic loop and the C terminus of α13.(C) Representative electron density in the synthetase site of monomer 1. GDP is highlighted in orange. The final 2mFo-DFc electron density map (1.0 σ) is overlaid as blue mesh.(D) Stereographic superposition between the synthetase site of Rel[Seq]1–385 (monomer 1) and the active site of pol β in the (pol β)·(gapped DNA)·(ddCTP) complex. The latter complex is rendered in gray shading with the exception of the primer 3′-terminal nucleotide (orange), ddCTP (cyan), and the two Mg^2+ ions (dark blue). Rel[Seq]1–385 and its GDP ligand are colored according to (A). The putative catalytic carboxylates of Rel[Seq], Asp264 and Glu323, are N terminally frameshifted by two residues relative to their pol β counterparts (indicated by black arrows).

The above figures are reprinted by permission from Cell Press: Cell (2004, 117, 57-68) copyright 2004.

Figures were selected by an automated process.