PDBsum entry 4mim

Ligase

PDB id: 4mim

Contents

Protein chains

595 a.a.

Ligands

BPV ×4

GOL ×2

Metals

_MG ×4

_ZN ×4

_CL ×4

Waters ×136

PDB id:

4mim

Name:

Ligase

Title:

Structure of the carboxyl transferase domain from rhizobium etli pyruvate carboxylase with 3-bromopyruvate

Structure:

Pyruvate carboxylase. Chain: a, b, c, d. Fragment: carboxyl transferase domain, unp residues 465-1067. Engineered: yes

Source:

Rhizobium etli. Organism_taxid: 347834. Strain: cfn 42. Gene: pyc, rhe_ch04002. Expressed in: escherichia coli. Expression_system_taxid: 469008.

Resolution:

2.65Å R-factor: 0.189 R-free: 0.235

Authors:

A.D.Lietzan,M.St.Maurice

Key ref:

A.D.Lietzan and M.St Maurice (2013). Insights into the carboxyltransferase reaction of pyruvate carboxylase from the structures of bound product and intermediate analogs. Biochem Biophys Res Commun, 441, 377-382. PubMed id: 24157795 DOI: 10.1016/j.bbrc.2013.10.066

Date:

01-Sep-13 Release date: 13-Aug-14

Protein chains

Q2K340  (Q2K340_RHIEC) -  Pyruvate carboxylase from Rhizobium etli (strain CFN 42 / ATCC 51251)

Seq: 1154 a.a.

Struc: 595 a.a.*

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

Enzyme reactions

• Enzyme class: E.C.6.4.1.1 - pyruvate carboxylase.
• Reaction: hydrogencarbonate + pyruvate + ATP = oxaloacetate + ADP + phosphate + H⁺

hydrogencarbonate + pyruvate + ATP (Bound ligand (Het Group name = BPV) matches with 85.71% similarity) = oxaloacetate + ADP + phosphate + H(+)

• Cofactor: Biotin; Mn(2+) or Zn(2+)

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

reference

DOI no: 10.1016/j.bbrc.2013.10.066

Biochem Biophys Res Commun 441:377-382 (2013)

PubMed id: 24157795

Insights into the carboxyltransferase reaction of pyruvate carboxylase from the structures of bound product and intermediate analogs.

A.D.Lietzan, M.St Maurice.

ABSTRACT

Pyruvate carboxylase (PC) is a biotin-dependent enzyme that catalyzes the MgATP- and bicarbonate-dependent carboxylation of pyruvate to oxaloacetate, an important anaplerotic reaction in central metabolism. The carboxyltransferase (CT) domain of PC catalyzes the transfer of a carboxyl group from carboxybiotin to the accepting substrate, pyruvate. It has been hypothesized that the reactive enolpyruvate intermediate is stabilized through a bidentate interaction with the metal ion in the CT domain active site. Whereas bidentate ligands are commonly observed in enzymes catalyzing reactions proceeding through an enolpyruvate intermediate, no bidentate interaction has yet been observed in the CT domain of PC. Here, we report three X-ray crystal structures of the Rhizobium etli PC CT domain with the bound inhibitors oxalate, 3-hydroxypyruvate, and 3-bromopyruvate. Oxalate, a stereoelectronic mimic of the enolpyruvate intermediate, does not interact directly with the metal ion. Instead, oxalate is buried in a pocket formed by several positively charged amino acid residues and the metal ion. Furthermore, both 3-hydroxypyruvate and 3-bromopyruvate, analogs of the reaction product oxaloacetate, bind in an identical manner to oxalate suggesting that the substrate maintains its orientation in the active site throughout catalysis. Together, these structures indicate that the substrates, products and intermediates in the PC-catalyzed reaction are not oriented in the active site as previously assumed. The absence of a bidentate interaction with the active site metal appears to be a unique mechanistic feature among the small group of biotin-dependent enzymes that act on α-keto acid substrates.