PDBsum entry 4j5n

Hydrolase

PDB id: 4j5n

Contents

Protein chain

322 a.a.

Metals

_CO ×2

Waters ×210

PDB id:

4j5n

Name:

Hydrolase

Title:

Crystal structure of a deinococcus radiodurans pte-like lactonase (drpll) mutant y28l/d71n/e101g/e179d/v235l/l270m

Structure:

Phosphotriesterase, putative. Chain: a. Engineered: yes. Mutation: yes

Source:

Deinococcus radiodurans. Organism_taxid: 1299. Gene: dr0930 (gi 15805954), dr_0930. Expressed in: escherichia coli. Expression_system_taxid: 469008.

Resolution:

2.05Å R-factor: 0.235 R-free: 0.261

Authors:

M.M.Meier,C.Rajendran,C.Malisi,N.G.Fox,C.Xu,S.Schlee,D.P.Barondeau, B.Hocker,R.Sterner,F.M.Raushel

Key ref:

M.M.Meier et al. (2013). Molecular engineering of organophosphate hydrolysis activity from a weak promiscuous lactonase template. J Am Chem Soc, 135, 11670-11677. PubMed id: 23837603 DOI: 10.1021/ja405911h

Date:

08-Feb-13 Release date: 24-Jul-13

Protein chain

Q9RVU2  (Q9RVU2_DEIRA) -  Phosphotriesterase, putative from Deinococcus radiodurans (strain ATCC 13939 / DSM 20539 / JCM 16871 / CCUG 27074 / LMG 4051 / NBRC 15346 / NCIMB 9279 / VKM B-1422 / R1)

Seq: 323 a.a.

Struc: 322 a.a.*

* PDB and UniProt seqs differ at 7 residue positions (black crosses)

DOI no: 10.1021/ja405911h

J Am Chem Soc 135:11670-11677 (2013)

PubMed id: 23837603

Molecular engineering of organophosphate hydrolysis activity from a weak promiscuous lactonase template.

M.M.Meier, C.Rajendran, C.Malisi, N.G.Fox, C.Xu, S.Schlee, D.P.Barondeau, B.Höcker, R.Sterner, F.M.Raushel.

ABSTRACT

Rapid evolution of enzymes provides unique molecular insights into the remarkable adaptability of proteins and helps to elucidate the relationship between amino acid sequence, structure, and function. We interrogated the evolution of the phosphotriesterase from Pseudomonas diminuta (PdPTE), which hydrolyzes synthetic organophosphates with remarkable catalytic efficiency. PTE is thought to be an evolutionarily "young" enzyme, and it has been postulated that it has evolved from members of the phosphotriesterase-like lactonase (PLL) family that show promiscuous organophosphate-degrading activity. Starting from a weakly promiscuous PLL scaffold (Dr0930 from Deinococcus radiodurans ), we designed an extremely efficient organophosphate hydrolase (OPH) with broad substrate specificity using rational and random mutagenesis in combination with in vitro activity screening. The OPH activity for seven organophosphate substrates was simultaneously enhanced by up to 5 orders of magnitude, achieving absolute values of catalytic efficiencies up to 10(6) M(-1) s(-1). Structural and computational analyses identified the molecular basis for the enhanced OPH activity of the engineered PLL variants and demonstrated that OPH catalysis in PdPTE and the engineered PLL differ significantly in the mode of substrate binding.