PDBsum entry: 1b4s

Phosphotransferase

PDB id: 1b4s

Contents

Protein chains

150 a.a. *

Ligands

PO4 ×3

ADP ×3

Metals

_MG ×3

Waters ×108

* Residue conservation analysis

PDB id:

1b4s

Name:

Phosphotransferase

Title:

Structure of nucleoside diphosphate kinase h122g mutant

Structure:

Nucleoside diphosphate kinase. Chain: a, b, c. Synonym: ndpk. Engineered: yes. Mutation: yes

Source:

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

Biol. unit:

Homo-Hexamer (from PDB file)

Resolution:

2.50Å R-factor: 0.203 R-free: 0.305

Authors:

P.Meyer,J.Janin

Key ref:

S.J.Admiraal et al. (1999). Nucleophilic activation by positioning in phosphoryl transfer catalyzed by nucleoside diphosphate kinase. Biochemistry, 38, 4701-4711. PubMed id: 10200157 DOI: 10.1021/bi9827565

Date:

28-Dec-98 Release date: 15-Jun-99

Protein chains

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

ATP + nucleoside diphosphate = ADP (Bound ligand (Het Group name = ADP) corresponds exactly) + 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 (11 terms)
• Biochemical function: nucleotide binding (6 terms)

reference

DOI no: 10.1021/bi9827565

Biochemistry 38:4701-4711 (1999)

PubMed id: 10200157

Nucleophilic activation by positioning in phosphoryl transfer catalyzed by nucleoside diphosphate kinase.

S.J.Admiraal, B.Schneider, P.Meyer, J.Janin, M.Véron, D.Deville-Bonne, D.Herschlag.

ABSTRACT

The nonenzymatic reaction of ATP with a nucleophile to generate ADP and a phosphorylated product proceeds via a dissociative transition state with little bond formation to the nucleophile. Consideration of the dissociative nature of the nonenzymatic transition state leads to the following question: To what extent can the nucleophile be activated in enzymatic phosphoryl transfer? We have addressed this question for the NDP kinase reaction. A mutant form of the enzyme lacking the nucleophilic histidine (H122G) can be chemically rescued for ATP attack by imidazole or other exogenous small nucleophiles. The ATP reaction is 50-fold faster with the wild-type enzyme, which has an imidazole nucleophile positioned for reaction by a covalent bond, than with H122G, which employs a noncovalently bound imidazole nucleophile [(kcat/KM)ATP]. Further, a 4-fold advantage for imidazole positioned in the nucleophile binding pocket created by the mutation is suggested from comparison of the reaction of H122G and ATP with an imidazole versus a water nucleophile, after correction for the intrinsic reactivities of imidazole and water toward ATP in solution. X-ray structural analysis shows no detectable rearrangement of the residues surrounding His 122 upon mutation to Gly 122. The overall rate effect of approximately 10(2)-fold for the covalent imidazole nucleophile relative to water is therefore attributed to positioning of the nucleophile with respect to the reactive phosphoryl group. This is underscored by the more deleterious effect of replacing ATP with AlphaTauPgammaS in the wild-type reaction than in the imidazole-rescued mutant reaction, as follows. For the wild-type, AlphaTauPgammaS presumably disrupts positioning between nucleophile and substrate, resulting in a large thio effect of 300-fold, whereas precise alignment is already disrupted in the mutant because there is no covalent bond to the nucleophile, resulting in a smaller thio effect of 10-fold. In summary, the results suggest a catalytic role for activation of the nucleophile by positioning in phosphoryl transfer catalyzed by NDP kinase.