PDBsum entry 4zwu

Hydrolase

PDB id: 4zwu

Contents

Protein chains

435 a.a.

Ligands

GOA

GOL

Metals

_MN ×6

_BA ×2

Waters ×470

PDB id:

4zwu

Name:

Hydrolase

Title:

Crystal structure of organophosphate anhydrolase/prolidase mutant y212f, v342l, i215y

Structure:

Organophosphate anhydrolase/prolidase. Chain: a, b. Synonym: x-pro dipeptidase,dfpase,imidodipeptidase,organophosphorus acid anhydrolase 2,opaa-2,paraoxon hydrolase,phosphotriesterase, proline dipeptidase,prolidase. Engineered: yes

Source:

Alteromonas sp.. Organism_taxid: 232. Gene: pepq, opaa. Expressed in: escherichia coli. Expression_system_taxid: 562.

Resolution:

2.20Å R-factor: 0.172 R-free: 0.219

Authors:

C.M.Daczkowski,S.D.Pegan,S.P.Harvey

Key ref:

C.M.Daczkowski et al. (2015). Engineering the Organophosphorus Acid Anhydrolase Enzyme for Increased Catalytic Efficiency and Broadened Stereospecificity on Russian VX. Biochemistry, 54, 6423-6433. PubMed id: 26418828 DOI: 10.1021/acs.biochem.5b00624

Date:

19-May-15 Release date: 14-Oct-15

Protein chains

Q44238  (PEPQ_ALTSX) -  Xaa-Pro dipeptidase from Alteromonas sp

Seq: 517 a.a.

Struc: 435 a.a.*

* PDB and UniProt seqs differ at 9 residue positions (black crosses)

Enzyme reactions

• Enzyme class 1: E.C.3.1.8.1 - aryldialkylphosphatase.
• Reaction: An aryl dialkyl phosphate + H2O = dialkyl phosphate + an aryl alcohol
• Cofactor: Divalent cation
• Enzyme class 2: E.C.3.4.13.9 - Xaa-Pro dipeptidase.
• Reaction: Xaa-L-Pro dipeptide + H2O = an L-alpha-amino acid + L-proline
• Cofactor: Mn(2+)
• Enzyme class 3: E.C.3.8.2.2 - diisopropyl-fluorophosphatase.
• Reaction: diisopropyl fluorophosphate + H2O = diisopropyl phosphate + fluoride + 2 H⁺

diisopropyl fluorophosphate + H2O = diisopropyl phosphate + fluoride (Bound ligand (Het Group name = GOA) matches with 44.44% similarity) + 2 × H(+)

Note, where more than one E.C. class is given (as above), each may correspond to a different protein domain or, in the case of polyprotein precursors, to a different mature protein.

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

reference

DOI no: 10.1021/acs.biochem.5b00624

Biochemistry 54:6423-6433 (2015)

PubMed id: 26418828

Engineering the Organophosphorus Acid Anhydrolase Enzyme for Increased Catalytic Efficiency and Broadened Stereospecificity on Russian VX.

C.M.Daczkowski, S.D.Pegan, S.P.Harvey.

ABSTRACT

The enzyme organophosphorus acid anhydrolase (OPAA), from Alteromonas sp. JD6.5, has been shown to rapidly catalyze the hydrolysis of a number of toxic organophosphorus compounds, including several G-type chemical nerve agents. The enzyme was cloned into Escherichia coli and can be produced up to approximately 50% of cellular protein. There have been no previous reports of OPAA activity on VR {Russian VX, O-isobutyl S-[2-(diethylamino)ethyl] methylphosphonothioate}, and our studies reported here show that wild-type OPAA has poor catalytic efficacy toward VR. However, via application of a structurally aided protein engineering approach, significant improvements in catalytic efficiency were realized via optimization of the small pocket within the OPAA's substrate-binding site. This optimization involved alterations at only three amino acid sites resulting in a 30-fold increase in catalytic efficiency toward racemic VR, with a strong stereospecificity toward the P(+) enantiomer. X-ray structures of this mutant as well as one of its predecessors provide potential structural rationales for their effect on the OPAA active site. Additionally, a fourth mutation at a site near the small pocket was found to relax the stereospecificity of the OPAA enzyme. Thus, it allows the altered enzyme to effectively process both VR enantiomers and should be a useful genetic background in which to seek further improvements in OPAA VR activity.