PDBsum entry 7clz

Hydrolase

PDB id: 7clz

Contents

Protein chains

293 a.a.

Ligands

DY9 ×3

MLT ×4

BO3 ×2

Metals

_NA ×2

Waters ×354

PDB id:

7clz

Name:

Hydrolase

Title:

Crystal structure of alp1u w187f/y247f in complex with fluostatin c

Structure:

Putative hydrolase. Chain: a, b, c, d. Synonym: alp1u. Engineered: yes. Mutation: yes

Source:

Streptomyces ambofaciens (strain atcc 23877 / 3486 / dsm 40053 / jcm 4204 / nbrc 12836 / nrrl b-2516). Organism_taxid: 278992. Gene: samt0137, samt0138. Expressed in: escherichia coli. Expression_system_taxid: 562

Resolution:

2.10Å R-factor: 0.196 R-free: 0.247

Authors:

L.Zhang,B.C.De

Key ref:

L.Zhang et al. (2020). Mutation of an atypical oxirane oxyanion hole improves regioselectivity of the α/β-fold epoxide hydrolase Alp1U. J Biol Chem, 295, 16987-16997. PubMed id: 33004437 DOI: 10.1074/jbc.RA120.015563

Date:

22-Jul-20 Release date: 07-Oct-20

Protein chains

A0A0K2AJY3  (A0A0K2AJY3_STRAM) -  Putative hydrolase from Streptomyces ambofaciens (strain ATCC 23877 / 3486 / DSM 40053 / JCM 4204 / NBRC 12836 / NRRL B-2516)

Seq: 326 a.a.

Struc: 293 a.a.*

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

DOI no: 10.1074/jbc.RA120.015563

J Biol Chem 295:16987-16997 (2020)

PubMed id: 33004437

Mutation of an atypical oxirane oxyanion hole improves regioselectivity of the α/β-fold epoxide hydrolase Alp1U.

L.Zhang, B.C.De, W.Zhang, A.Mándi, Z.Fang, C.Yang, Y.Zhu, T.Kurtán, C.Zhang.

ABSTRACT

Epoxide hydrolases (EHs) have been characterized and engineered as biocatalysts that convert epoxides to valuable chiral vicinal diol precursors of drugs and bioactive compounds. Nonetheless, the regioselectivity control of the epoxide ring opening by EHs remains challenging. Alp1U is an α/β-fold EH that exhibits poor regioselectivity in the epoxide hydrolysis of fluostatin C (compound 1) and produces a pair of stereoisomers. Herein, we established the absolute configuration of the two stereoisomeric products and determined the crystal structure of Alp1U. A Trp-186/Trp-187/Tyr-247 oxirane oxygen hole was identified in Alp1U that replaced the canonical Tyr/Tyr pair in α/β-EHs. Mutation of residues in the atypical oxirane oxygen hole of Alp1U improved the regioselectivity for epoxide hydrolysis on 1. The single site Y247F mutation led to highly regioselective (98%) attack at C-3 of 1, whereas the double mutation W187F/Y247F resulted in regioselective (94%) nucleophilic attack at C-2. Furthermore, single-crystal X-ray structures of the two regioselective Alp1U variants in complex with 1 were determined. These findings allowed insights into the reaction details of Alp1U and provided a new approach for engineering regioselective epoxide hydrolases.