PDBsum entry: 1djo

Hydrolase/hydrolase inhibitor

PDB id: 1djo

Contents

Protein chains

330 a.a. *

Ligands

CAB ×2

* Residue conservation analysis

PDB id:

1djo

Name:

Hydrolase/hydrolase inhibitor

Title:

Crystal structure of pseudomonas 7a glutaminase-asparaginase inhibitor donv covalently bound in the active site

Structure:

Glutaminase-asparaginase. Chain: a, b. Ec: 3.5.1.38

Source:

Pseudomonas sp.. Organism_taxid: 65406. Strain: 7a

Biol. unit:

Tetramer (from PDB file)

Resolution:

2.00Å R-factor: 0.214 R-free: 0.238

Authors:

E.Ortlund,M.W.Lacount,K.Lewinski,L.Lebioda

Key ref:

E.Ortlund et al. (2000). Reactions of Pseudomonas 7A glutaminase-asparaginase with diazo analogues of glutamine and asparagine result in unexpected covalent inhibitions and suggests an unusual catalytic triad Thr-Tyr-Glu. Biochemistry, 39, 1199-1204. PubMed id: 10684596 DOI: 10.1021/bi991797d

Date:

03-Dec-99 Release date: 24-Jan-00

Protein chains

P10182  (ASPQ_PSES7) -  Glutaminase-asparaginase

Seq: 337 a.a.

Struc: 330 a.a.*

* PDB and UniProt seqs differ at 12 residue positions (black crosses)

Enzyme reactions

• Enzyme class: E.C.3.5.1.38 - Glutamin-(asparagin-)ase.
• Reaction:
  1. L-glutamine + H₂O = L-glutamate + NH₃
  2. L-asparagine + H₂O = L-aspartate + NH₃

L-glutamine (Bound ligand (Het Group name = CAB) matches with 90.00% similarity) + H(2)O = L-glutamate + NH(3)

L-asparagine (Bound ligand (Het Group name = CAB) matches with 63.64% similarity) + H(2)O = L-aspartate + NH(3)

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

 Gene Ontology (GO) functional annotation 

• Cellular component: periplasmic space (1 term)
• Biological process: cellular amino acid metabolic process (2 terms)
• Biochemical function: hydrolase activity (3 terms)

reference

DOI no: 10.1021/bi991797d

Biochemistry 39:1199-1204 (2000)

PubMed id: 10684596

Reactions of Pseudomonas 7A glutaminase-asparaginase with diazo analogues of glutamine and asparagine result in unexpected covalent inhibitions and suggests an unusual catalytic triad Thr-Tyr-Glu.

E.Ortlund, M.W.Lacount, K.Lewinski, L.Lebioda.

ABSTRACT

Pseudomonas 7A glutaminase-asparaginase (PGA) catalyzes the hydrolysis of D and L isomers of glutamine and asparagine. Crystals of PGA were reacted with diazo analogues of glutamine (6-diazo-5-oxo-L-norleucine, DON) and asparagine (5-diazo-4-oxo-L-norvaline, DONV), which are known inhibitors of the enzyme. The derivatized crystals remained isomorphous to native PGA crystals. Their structures were refined to crystallographic R = 0.20 and R(free) = 0.24 for PGA-DON and R = 0.19 and R = 0.23 for PGA-DONV. Difference Fourier electron density maps clearly showed that both DON and DONV inactivate PGA through covalent inhibition. Continuous electron density connecting the inhibitor to both Thr20 and Tyr34 of the flexible loop was observed providing strong evidence that Thr20 is the primary catalytic nucleophile and that Tyr34 plays an important role in catalysis as well. The unexpected covalent binding observed in the PGA-DON and PGA-DONV complexes shows that a secondary reaction involving the formation of a Tyr34-inhibitor bond takes place with concomitant inactivation of PGA. The predicted covalent linkage is not seen, however, suggesting an alternative method of inhibition not yet seen for these diazo analogues. These surprising results give insight as to the role of the flexible loop Thr and Tyr in the catalytic mechanism.