PDBsum entry 1d09

Transferase

PDB id: 1d09

Contents

Protein chains

310 a.a. *

153 a.a. *

Ligands

PAL ×2

Metals

_ZN ×2

Waters ×617

* Residue conservation analysis

PDB id:

1d09

Name:

Transferase

Title:

Aspartate transcarbamoylase complexed with n-phosphonacetyl-l- aspartate (pala)

Structure:

Aspartate carbamoyltransferase catalytic chain. Chain: a, c. Engineered: yes. Other_details: complexed with n-phosphonacetyl-l-aspartate. Aspartate carbamoyltransferase regulatory chain. Chain: b, d. Engineered: yes

Source:

Escherichia coli. Organism_taxid: 562. Gene: natural pyrb promotor. Expressed in: escherichia coli. Expression_system_taxid: 562. Inserted into puc119. Gene: pyri. Inserted into puc119

Biol. unit:

Dodecamer (from PDB file)

Resolution:

2.10Å R-factor: 0.203 R-free: 0.234

Authors:

L.Jin,B.Stec,W.N.Lipscomb,E.R.Kantrowitz

Key ref:

L.Jin et al. (1999). Insights into the mechanisms of catalysis and heterotropic regulation of Escherichia coli aspartate transcarbamoylase based upon a structure of the enzyme complexed with the bisubstrate analogue N-phosphonacetyl-L-aspartate at 2.1 A. Proteins, 37, 729-742. PubMed id: 10651286 DOI: 10.1002/(SICI)1097-0134(19991201)37:4<729::AID-PROT21>3.3.CO;2-6

Date:

09-Sep-99 Release date: 28-Jan-00

Protein chains

P0A786  (PYRB_ECOLI) -  Aspartate carbamoyltransferase catalytic subunit from Escherichia coli (strain K12)

Seq: 311 a.a.

Struc: 310 a.a.*

Protein chains

P0A7F3  (PYRI_ECOLI) -  Aspartate carbamoyltransferase regulatory chain from Escherichia coli (strain K12)

Seq: 153 a.a.

Struc: 153 a.a.

* PDB and UniProt seqs differ at 2 residue positions (black crosses)

Enzyme reactions

• Enzyme class: Chains A, C: E.C.2.1.3.2 - aspartate carbamoyltransferase.
• Pathway: Pyrimidine Biosynthesis
• Reaction: carbamoyl phosphate + L-aspartate = N-carbamoyl-L-aspartate + phosphate + H⁺

carbamoyl phosphate + L-aspartate = N-carbamoyl-L-aspartate + phosphate (Bound ligand (Het Group name = PAL) matches with 64.71% similarity) + H(+)

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

reference

DOI no: 10.1002/(SICI)1097-0134(19991201)37:4<729::AID-PROT21>3.3.CO;2-6

Proteins 37:729-742 (1999)

PubMed id: 10651286

Insights into the mechanisms of catalysis and heterotropic regulation of Escherichia coli aspartate transcarbamoylase based upon a structure of the enzyme complexed with the bisubstrate analogue N-phosphonacetyl-L-aspartate at 2.1 A.

L.Jin, B.Stec, W.N.Lipscomb, E.R.Kantrowitz.

ABSTRACT

A high-resolution structure of Escherichia coli aspartate transcarbamoylase has been determined to 2.1 A; resolution in the presence of the bisubstrate analog N-phosphonacetyl-L-aspartate (PALA). The structure was refined to a free R-factor of 23.4% and a working R-factor of 20.3%. The PALA molecule is completely saturated with interactions to side chain and backbone groups in the active site, including two interactions that are contributed from the 80s loop of the adjacent catalytic chain. The charge neutralization of the bound PALA molecule (and presumably the substrates as well) induced by the electrostatic field of the highly positively charged active site is an important factor in the high binding affinity of PALA and must be important for catalysis. The higher-resolution structure reported here departs in a number of ways from the previously determined structure at lower resolution. These modifications include alterations in the backbone conformation of the C-terminal of the catalytic chains, the N- and C-termini of the regulatory chains, and two loops of the regulatory chain. The high-resolution of this structure has allowed a more detailed description of the binding of PALA to the active site of the enzyme and has allowed a detailed model of the tetrahedral intermediate to be constructed. This model becomes the basis of a description of the catalytic mechanism of the transcarbamoylase reaction. The R-structural state of the enzyme-PALA complex is an excellent representation of the form of the enzyme that occurs at the moment in the catalytic cycle when the tetrahedral intermediate is formed. Finally, improved electron density in the N-terminal region of the regulatory chain (residues 1 to 7) has allowed tracing of the entire regulatory chain. The N-terminal segments of the R1 and R6 chains are located in close proximity to each other and to the regulatory site. This portion of the molecule may be involved in the observed asymmetry between the regulatory binding sites as well as in the heterotropic response of the enzyme.

Selected figure(s)

Figure 1.
Figure 1. Schematic diagram of the PALA binding site in the C1 chain of aspartate transcarbamoylase. Shown are all of the residues that have hydrogen bonding interactions (dashed lines) with PALA. The only substantial difference between the C1 and C6 active sites is a reorientation of the side chain of Arg54.

Figure 6.
Figure 6. Stereo view of the C1 catalytic chain of aspartate transcarbamoylase with the tetrahedral intermediate modeled into the active site. Shown are all the side chains that make direct contact with the intermediate from the C1 chain as well as Ser80 and Lys84 from the adjacent catalytic chain. Hydrogen bonds are shown as dashed lines.

The above figures are reprinted by permission from John Wiley & Sons, Inc.: Proteins (1999, 37, 729-742) copyright 1999.

Figures were selected by an automated process.