PDBsum entry: 1ncl

Transferase

PDB id: 1ncl

Contents

Protein chain

150 a.a. *

Waters ×91

* Residue conservation analysis

PDB id:

1ncl

Name:

Transferase

Title:

Thermal stability of hexameric and tetrameric nucleoside, di kinases

Structure:

Nucleoside diphosphate kinase. Chain: a. Synonym: ndk, ndp kinase. Engineered: yes. Mutation: yes

Source:

Dictyostelium discoideum. Organism_taxid: 44689. Expressed in: escherichia coli. Expression_system_taxid: 562

Biol. unit:

Hexamer (from PQS)

Resolution:

2.20Å R-factor: 0.181

Authors:

J.Janin,S.Morera,I.Lascu

Key ref:

A.Giartosio et al. (1996). Thermal stability of hexameric and tetrameric nucleoside diphosphate kinases. Effect of subunit interaction. J Biol Chem, 271, 17845-17851. PubMed id: 8663370 DOI: 10.1074/jbc.271.30.17845

Date:

22-Mar-96 Release date: 08-Nov-96

Protein chain

P22887  (NDKC_DICDI) -  Nucleoside diphosphate kinase, cytosolic

Seq: 155 a.a.

Struc: 150 a.a.*

* PDB and UniProt seqs differ at 1 residue position (black cross)

Enzyme reactions

• Enzyme class: E.C.2.7.4.6 - Nucleoside-diphosphate kinase.
• Reaction: ATP + nucleoside diphosphate = ADP + nucleoside triphosphate

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

 Gene Ontology (GO) functional annotation 

• Cellular component: plasma membrane (6 terms)
• Biological process: cytoskeleton organization (13 terms)
• Biochemical function: nucleotide binding (6 terms)

reference

DOI no: 10.1074/jbc.271.30.17845

J Biol Chem 271:17845-17851 (1996)

PubMed id: 8663370

Thermal stability of hexameric and tetrameric nucleoside diphosphate kinases. Effect of subunit interaction.

A.Giartosio, M.Erent, L.Cervoni, S.Moréra, J.Janin, M.Konrad, I.Lascu.

ABSTRACT

The eukaryotic nucleoside diphosphate (NDP) kinases are hexamers, while the bacterial NDP kinases are tetramers made of small, single domain subunits. These enzymes represent an ideal model for studying the effect of subunit interaction on protein stability. The thermostability of NDP kinases of each class was studied by differential scanning calorimetry and biochemical methods. The hexameric NDP kinase from Dictyostelium discoideum displays one single, irreversible differential scanning calorimetry peak (Tm 62 degrees C) over a broad protein concentration, indicating a single step denaturation. The thermal stability of the protein was increased by ADP. The P105G substitution, which affects a loop implicated in subunit contacts, yields a protein that reversibly dissociates to folded monomers at 38 degrees C before the irreversible denaturation occurs (Tm 47 degrees C). ADP delays the dissociation, but does not change the Tm. These data indicate a "coupling" of the quaternary structure with the tertiary structure in the wild-type, but not in the mutated protein. We describe the x-ray structure of the P105G mutant at 2.2-A resolution. It is very similar to that of the wild-type protein. Therefore, a minimal change in the structure leads to a dramatic change of protein thermostability. The NDP kinase from Escherichia coli behaves like the P105G mutant of the Dictyostelium NDP kinase. The detailed study of their thermostability is important, since biological effects of thermolabile NDP kinases have been described in several organisms.

Selected figure(s)

Figure 1.
Fig. 1. Structure of the P105G protein. A, one-half of the NDP kinase hexamer viewed along the 3-fold axis. The boxed region is detailed in the stereo pair (B): the Kpn loops in the wild-type (empty bonds) and P105G (full bonds) proteins. Residues 100-105 are in ball-and-sticks. In the mutant, the Pro-105 side chain is replaced by water molecule W800. In both proteins, water molecule W803 hydrogen bonds to the carbonyl of Gly-106 and to its symmetry-related counterparts. The figure was created using MolScript (Kraulis, 1991[ref-arrow.gif]).

Figure 2.
Fig. 2. Heat inactivation of Dictyostelium NDP kinase. The wild-type ( ) and the P105G mutant ( ) at 1 mg/ml in 50 mM Hepes, pH 7.5, were heated at a rate of 60 °C/h. The residual activity was measured in the standard assay (for details see ``Materials and Methods'').

The above figures are reprinted by permission from the ASBMB: J Biol Chem (1996, 271, 17845-17851) copyright 1996.

Figures were selected by an automated process.