PDBsum entry 2bnf

Transferase

PDB id: 2bnf

Contents

Protein chains

236 a.a. *

Ligands

UTP ×2

GOL ×7

Waters ×74

* Residue conservation analysis

PDB id:

2bnf

Name:

Transferase

Title:

The structure of e. Coli ump kinase in complex with utp

Structure:

Uridylate kinase. Chain: a, b. Synonym: ump kinase, uk, uridine monophosphate kinase. Engineered: yes. Mutation: yes

Source:

Escherichia coli. Organism_taxid: 83333. Strain: k12. Expressed in: escherichia coli. Expression_system_taxid: 562

Biol. unit:

Hexamer (from PDB file)

Resolution:

2.45Å R-factor: 0.184 R-free: 0.231

Authors:

P.Briozzo,C.Evrin,P.Meyer,L.Assairi,N.Joly,O.Barzu,A.M.Gilles

Key ref:

P.Briozzo et al. (2005). Structure of Escherichia coli UMP kinase differs from that of other nucleoside monophosphate kinases and sheds new light on enzyme regulation. J Biol Chem, 280, 25533-25540. PubMed id: 15857829 DOI: 10.1074/jbc.M501849200

Date:

23-Mar-05 Release date: 27-Apr-05

Protein chains

P0A7E9  (PYRH_ECOLI) -  Uridylate kinase from Escherichia coli (strain K12)

Seq: 241 a.a.

Struc: 236 a.a.*

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

Enzyme reactions

• Enzyme class: E.C.2.7.4.22 - Ump kinase.
• Reaction: UMP + ATP = UDP + ADP

UMP + ATP = UDP + ADP (Bound ligand (Het Group name = UTP) matches with 86.21% similarity)

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

reference

DOI no: 10.1074/jbc.M501849200

J Biol Chem 280:25533-25540 (2005)

PubMed id: 15857829

Structure of Escherichia coli UMP kinase differs from that of other nucleoside monophosphate kinases and sheds new light on enzyme regulation.

P.Briozzo, C.Evrin, P.Meyer, L.Assairi, N.Joly, O.Barzu, A.M.Gilles.

ABSTRACT

Bacterial UMP kinases are essential enzymes involved in the multistep synthesis of nucleoside triphosphates. They are hexamers regulated by the allosteric activator GTP and inhibited by UTP. We solved the crystal structure of Escherichia coli UMP kinase bound to the UMP substrate (2.3 A resolution), the UDP product (2.6 A), or UTP (2.45 A). The monomer fold, unrelated to that of other nucleoside monophosphate kinases, belongs to the carbamate kinase-like superfamily. However, the phosphate acceptor binding cleft and subunit assembly are characteristic of UMP kinase. Interactions with UMP explain the high specificity for this natural substrate. UTP, previously described as an allosteric inhibitor, was unexpectedly found in the phosphate acceptor site, suggesting that it acts as a competitive inhibitor. Site-directed mutagenesis of residues Thr-138 and Asn-140, involved in both uracil recognition and active site interaction within the hexamer, decreased the activation by GTP and inhibition by UTP. These experiments suggest a cross-talk mechanism between enzyme subunits involved in cooperative binding at the phosphate acceptor site and in allosteric regulation by GTP. As bacterial UMP kinases have no counterpart in eukaryotes, the information provided here could help the design of new antibiotics.

Selected figure(s)

Figure 2.
FIG. 2. Overall fold and quaternary structure. A, ribbon representation of the UMPKeco monomer fold (using the UMP-containing complex). -strands (yellow) and helices (blue, except 3, pink) are numbered. A stick model of UMP is shown in magenta. The cross-talk loop (green) is labeled CT. The loops are smoothed for clarity. A gray line connects residues that delimit a segment in which no clear density can be seen. B, the dimer constituted by the two molecules of an asymmetric unit. View along the non-crystallographic two-fold axis (indicated by a black ellipse symbol). The blue subunit orientation is close to that in panel A. C, ribbon representation of the hexamer viewed along the three-fold crystallographic axis (indicated by a black triangle). A particular color is used for each subunit. For the blue and green dimer, 3 helices are pink. Corey-Pauling-Koltun space-filling models of UMP are magenta. The three non-crystallographic two-fold axes are shown by dotted lines; they are perpendicular to the three-fold axis. D, magnification of the dimer-dimer interface emphasizing the 2 residues (shown in sticks and labeled) from the cross-talk loop that interact both with their homologues from the facing dimer and with UMP (stick model with carbon atoms in magenta). Hydrogen bonds are shown as red dots. -helices are transparized for clarity. The figure was drawn with PyMOL, Version 0.97 (38).

Figure 3.
FIG. 3. Comparison with NAGK structure. A, superposition of the ribbon representations of the monomers of UMPKeco (blue, with a stick model of UMP in cyan) and NAGK (yellow, with N-acetyl glutamine and ADPNP in red; the helix homologous to 3 is pink). The cross-talk loop of UMPKeco is green, the flexible loops close to ADPNP are magenta, and the extra -hairpins of NAGK are orange. The orientation is close to that in Fig. 2A but slightly modified to better see all ligands. B, the dimer of NAGK. The orientation is slightly different from that in Fig. 2B to give a better view of the interface.

The above figures are reprinted by permission from the ASBMB: J Biol Chem (2005, 280, 25533-25540) copyright 2005.

Figures were selected by the author.