PDBsum entry 2zma

Hydrolase

PDB id: 2zma

Contents

Protein chain

384 a.a. *

Ligands

SO4 ×5

ACA-ACA

MES

GOL ×5

Waters ×430

* Residue conservation analysis

PDB id:

2zma

Name:

Hydrolase

Title:

Crystal structure of 6-aminohexanoate-dimer hydrolase s112a/g181d/h266n/d370y mutant with substrate

Structure:

6-aminohexanoate-dimer hydrolase. Chain: a. Synonym: nylon oligomers-degrading enzyme eii, nylon oligomers- degrading enzyme eii'. Engineered: yes. Mutation: yes. Other_details: chimera of nylon oligomers-degrading enzyme eii (residues 1-21) and nylon oligomers-degrading enzyme eii' (residues 22-392)

Source:

Flavobacterium sp.. Organism_taxid: 239. Gene: nylb, nylb'. Expressed in: escherichia coli. Expression_system_taxid: 562.

Resolution:

1.51Å R-factor: 0.175 R-free: 0.190

Authors:

T.Ohki,N.Shibata,Y.Higuchi,M.Takeo,S.Negoro

Key ref:

Y.Kawashima et al. (2009). Molecular design of a nylon-6 byproduct-degrading enzyme from a carboxylesterase with a beta-lactamase fold. Febs J, 276, 2547-2556. PubMed id: 19476493

Date:

14-Apr-08 Release date: 07-Apr-09

Protein chain

P07061  (NYLB_PAEUR) -  6-aminohexanoate-dimer hydrolase from Paenarthrobacter ureafaciens

Seq: 392 a.a.

Struc: 384 a.a.*

Protein chain

P07062  (NYLB2_PAEUR) -  6-aminohexanoate-dimer hydrolase from Paenarthrobacter ureafaciens

Seq: 392 a.a.

Struc: 384 a.a.*

* PDB and UniProt seqs differ at 46 residue positions (black crosses)

Enzyme reactions

• Enzyme class: E.C.3.5.1.46 - 6-aminohexanoate-oligomer exohydrolase.
• Reaction:
  1. [N-(6-aminohexanoyl)](n) + H2O = [N-(6-aminohexanoyl)](n-1) + 6-aminohexanoate
  2. N-(6-aminohexanoyl)-6-aminohexanoate + H2O = 2 6-aminohexanoate

[N-(6-aminohexanoyl)](n) + H2O = [N-(6-aminohexanoyl)](n-1) (Bound ligand (Het Group name = ACA) corresponds exactly) + 6-aminohexanoate

N-(6-aminohexanoyl)-6-aminohexanoate + H2O = 2 × 6-aminohexanoate (Bound ligand (Het Group name = ACA) corresponds exactly)

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

reference

Febs J 276:2547-2556 (2009)

PubMed id: 19476493

Molecular design of a nylon-6 byproduct-degrading enzyme from a carboxylesterase with a beta-lactamase fold.

Y.Kawashima, T.Ohki, N.Shibata, Y.Higuchi, Y.Wakitani, Y.Matsuura, Y.Nakata, M.Takeo, D.Kato, S.Negoro.

ABSTRACT

A carboxylesterase with a beta-lactamase fold from Arthrobacter possesses a low level of hydrolytic activity (0.023 mumol.min(-1).mg(-1)) when acting on a 6-aminohexanoate linear dimer byproduct of the nylon-6 industry (Ald). G181D/H266N/D370Y triple mutations in the parental esterase increased the Ald-hydrolytic activity 160-fold. Kinetic studies showed that the triple mutant possesses higher affinity for the substrate Ald (K(m) = 2.0 mm) than the wild-type Ald hydrolase from Arthrobacter (K(m) = 21 mm). In addition, the k(cat)/K(m) of the mutant (1.58 s(-1).mm(-1)) was superior to that of the wild-type enzyme (0.43 s(-1).mm(-1)), demonstrating that the mutant efficiently converts the unnatural amide compounds even at low substrate concentrations, and potentially possesses an advantage for biotechnological applications. X-ray crystallographic analyses of the G181D/H266N/D370Y enzyme and the inactive S112A-mutant-Ald complex revealed that Ald binding induces rotation of Tyr370/His375, movement of the loop region (N167-V177), and flip-flop of Tyr170, resulting in the transition from open to closed forms. From the comparison of the three-dimensional structures of various mutant enzymes and site-directed mutagenesis at positions 266 and 370, we now conclude that Asn266 makes suitable contacts with Ald and improves the electrostatic environment at the N-terminal region of Ald cooperatively with Asp181, and that Tyr370 stabilizes Ald binding by hydrogen-bonding/hydrophobic interactions at the C-terminal region of Ald.