PDBsum entry 4j5h

Hydrolase/hydrolase substrate

PDB id: 4j5h

Contents

Protein chain

250 a.a.

Ligands

1K4

GOL ×8

Metals

_ZN ×2

Waters ×168

PDB id:

4j5h

Name:

Hydrolase/hydrolase substrate

Title:

Crystal structure of b. Thuringiensis aiia mutant f107w with n- decanoyl-l-homoserine bound at the active site

Structure:

N-acyl homoserine lactonase. Chain: a. Synonym: ahl-lactonase, homoserine lactone lactonase. Engineered: yes. Mutation: yes

Source:

Bacillus thuringiensis. Organism_taxid: 1428. Gene: aiia. Expressed in: escherichia coli. Expression_system_taxid: 469008.

Resolution:

1.45Å R-factor: 0.122 R-free: 0.151

Authors:

C.F.Liu,D.Liu,J.Momb,P.W.Thomas,A.Lajoie,G.A.Petsko,W.Fast,D.Ringe

Key ref:

C.F.Liu et al. (2013). A phenylalanine clamp controls substrate specificity in the quorum-quenching metallo-γ-lactonase from Bacillus thuringiensis. Biochemistry, 52, 1603-1610. PubMed id: 23387521 DOI: 10.1021/bi400050j

Date:

08-Feb-13 Release date: 26-Jun-13

Protein chain

A3FJ64  (AHLL_BACTU) -  N-acyl homoserine lactonase from Bacillus thuringiensis

Seq: 250 a.a.

Struc: 250 a.a.*

* PDB and UniProt seqs differ at 1 residue position (black cross)

Enzyme reactions

• Enzyme class: E.C.3.1.1.81 - quorum-quenching N-acyl-homoserine lactonase.
• Reaction: an N-acyl-L-homoserine lactone + H2O = an N-acyl-L-homoserine + H⁺

N-acyl-L-homoserine lactone + H2O = N-acyl-L-homoserine (Bound ligand (Het Group name = 1K4) matches with 50.00% similarity) + H(+)

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

Added reference

DOI no: 10.1021/bi400050j

Biochemistry 52:1603-1610 (2013)

PubMed id: 23387521

A phenylalanine clamp controls substrate specificity in the quorum-quenching metallo-γ-lactonase from Bacillus thuringiensis.

C.F.Liu, D.Liu, J.Momb, P.W.Thomas, A.Lajoie, G.A.Petsko, W.Fast, D.Ringe.

ABSTRACT

Autoinducer inactivator A (AiiA) is a metal-dependent N-acyl homoserine lactone hydrolase that displays broad substrate specificity but shows a preference for substrates with long N-acyl substitutions. Previously, crystal structures of AiiA in complex with the ring-opened product N-hexanoyl-l-homoserine revealed binding interactions near the metal center but did not identify a binding pocket for the N-acyl chains of longer substrates. Here we report the crystal structure of an AiiA mutant, F107W, determined in the presence and absence of N-decanoyl-l-homoserine. F107 is located in a hydrophobic cavity adjacent to the previously identified ligand binding pocket, and the F107W mutation results in the formation of an unexpected interaction with the ring-opened product. Notably, the structure reveals a previously unidentified hydrophobic binding pocket for the substrate's N-acyl chain. Two aromatic residues, F64 and F68, form a hydrophobic clamp, centered around the seventh carbon in the product-bound structure's decanoyl chain, making an interaction that would also be available for longer substrates, but not for shorter substrates. Steady-state kinetics using substrates of various lengths with AiiA bearing mutations at the hydrophobic clamp, including insertion of a redox-sensitive cysteine pair, confirms the importance of this hydrophobic feature for substrate preference. Identifying the specificity determinants of AiiA will aid the development of more selective quorum-quenching enzymes as tools and as potential therapeutics.