PDBsum entry 2f17

Transferase

PDB id: 2f17

Contents

Protein chains

255 a.a. *

Ligands

SO4 ×6

PYI ×2

AMP

EPE

Metals

_MG ×2

Waters ×132

* Residue conservation analysis

PDB id:

2f17

Name:

Transferase

Title:

Mouse thiamin pyrophosphokinase in a ternary complex with pyrithiamin pyrophosphate and amp at 2.5 angstrom

Structure:

Thiamin pyrophosphokinase 1. Chain: a, b. Synonym: thiamine pyrophosphokinase 1, mtpk1. Engineered: yes

Source:

Mus musculus. House mouse. Organism_taxid: 10090. Gene: tpk1. Expressed in: escherichia coli bl21. Expression_system_taxid: 511693.

Biol. unit:

Tetramer (from PQS)

Resolution:

2.50Å R-factor: 0.237 R-free: 0.252

Authors:

J.Y.Liu,D.E.Timm,T.D.Hurley

Key ref:

J.Y.Liu et al. (2006). Pyrithiamine as a substrate for thiamine pyrophosphokinase. J Biol Chem, 281, 6601-6607. PubMed id: 16365036 DOI: 10.1074/jbc.M510951200

Date:

14-Nov-05 Release date: 29-Nov-05

Protein chains

Q9R0M5  (TPK1_MOUSE) -  Thiamin pyrophosphokinase 1 from Mus musculus

Seq: 243 a.a.

Struc: 255 a.a.

Enzyme reactions

• Enzyme class: E.C.2.7.6.2 - thiamine diphosphokinase.
• Reaction: thiamine + ATP = thiamine diphosphate + AMP + H⁺

thiamine + ATP = thiamine diphosphate (Bound ligand (Het Group name = AMP) corresponds exactly) + AMP + H(+) (Bound ligand (Het Group name = PYI) matches with 82.76% similarity)

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

reference

DOI no: 10.1074/jbc.M510951200

J Biol Chem 281:6601-6607 (2006)

PubMed id: 16365036

Pyrithiamine as a substrate for thiamine pyrophosphokinase.

J.Y.Liu, D.E.Timm, T.D.Hurley.

ABSTRACT

Thiamine pyrophosphokinase transfers a pyrophosphate group from a nucleoside triphosphate, such as ATP, to the hydroxyl group of thiamine to produce thiamine pyrophosphate. Deficiencies in thiamine can result in the development of the neurological disorder Wernicke-Korsakoff Syndrome as well as the potentially fatal cardiovascular disease wet beriberi. Pyrithiamine is an inhibitor of thiamine metabolism that induces neurological symptoms similar to that of Wernicke-Korsakoff Syndrome in animals. However, the mechanism by which pyrithiamine interferes with cellular thiamine phosphoester homeostasis is not entirely clear. We used kinetic assays coupled with mass spectrometry of the reaction products and x-ray crystallography of an equilibrium reaction mixture of thiamine pyrophosphokinase, pyrithiamine, and Mg2+/ATP to elucidate the mechanism by which pyrithiamine inhibits the enzymatic production of thiamine pyrophosphate. Three lines of evidence support the ability of thiamine pyrophosphokinase to form pyrithiamine pyrophosphate. First, a coupled enzyme assay clearly demonstrated the ability of thiamine pyrophosphokinase to produce AMP when pyrithiamine was used as substrate. Second, an analysis of the reaction mixture by mass spectrometry directly identified pyrithiamine pyrophosphate in the reaction mixture. Last, the structure of thiamine pyrophosphokinase crystallized from an equilibrium substrate/product mixture shows clear electron density for pyrithiamine pyrophosphate bound in the enzyme active site. This structure also provides the first clear picture of the binding pocket for the nucleoside triphosphate and permits the first detailed understanding of the catalytic requirements for catalysis in this enzyme.

Selected figure(s)

Figure 1.
FIGURE 1. The chemical structures of thiamine, oxythiamine, and pyrithiamine.

Figure 2.
FIGURE 2. Electron density maps of pyrithiamine pyrophosphate and Mg^2+/AMP. A, omit 2F[o]-F[c] map of the bound PPP molecule contoured at 1 S.D. of the map. In the upper panel, a F[o]-F[c] map contoured at 3 S.D. shows the negative peak surrounding the sulfur atom of TPP after replacing PPP with TPP and performing a round of refinement in CNS. B, omit 2F[o]-F[c] map of Mg^2+/AMP bound to TPK contoured at 1 S.D. of the map.

The above figures are reprinted by permission from the ASBMB: J Biol Chem (2006, 281, 6601-6607) copyright 2006.

Figures were selected by the author.