PDBsum entry 2oqw

Hydrolase

PDB id: 2oqw

Contents

Protein chain

209 a.a. *

Waters ×85

* Residue conservation analysis

PDB id:

2oqw

Name:

Hydrolase

Title:

The crystal structure of sortase b from b.Anthracis in complex with aaek1

Structure:

Sortase b. Chain: a. Synonym: hypothetical protein. Engineered: yes

Source:

Bacillus anthracis str.. Organism_taxid: 198094. Strain: ames. Gene: ba_4783, bas4438, gbaa4783. Expressed in: escherichia coli. Expression_system_taxid: 562.

Resolution:

2.10Å R-factor: 0.190 R-free: 0.259

Authors:

R.Wu,R.Zhang,A.W.Marresso,O.Schneewind,A.Joachimiak

Key ref:

A.W.Maresso et al. (2007). Activation of inhibitors by sortase triggers irreversible modification of the active site. J Biol Chem, 282, 23129-23139. PubMed id: 17545669 DOI: 10.1074/jbc.M701857200

Date:

01-Feb-07 Release date: 19-Jun-07

Protein chain

A0A6L8PZR0  (A0A6L8PZR0_BACAN) -  SrtB family sortase from Bacillus anthracis

Seq: 254 a.a.

Struc: 209 a.a.*

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

DOI no: 10.1074/jbc.M701857200

J Biol Chem 282:23129-23139 (2007)

PubMed id: 17545669

Activation of inhibitors by sortase triggers irreversible modification of the active site.

A.W.Maresso, R.Wu, J.W.Kern, R.Zhang, D.Janik, D.M.Missiakas, M.E.Duban, A.Joachimiak, O.Schneewind.

ABSTRACT

Sortases anchor surface proteins to the cell wall of Gram-positive pathogens through recognition of specific motif sequences. Loss of sortase leads to large reductions in virulence, which identifies sortase as a target for the development of antibacterials. By screening 135,625 small molecules for inhibition, we report here that aryl (beta-amino)ethyl ketones inhibit sortase enzymes from staphylococci and bacilli. Inhibition of sortases occurs through an irreversible, covalent modification of their active site cysteine. Sortases specifically activate this class of molecules via beta-elimination, generating a reactive olefin intermediate that covalently modifies the cysteine thiol. Analysis of the three-dimensional structure of Bacillus anthracis sortase B with and without inhibitor provides insights into the mechanism of inhibition and reveals binding pockets that can be exploited for drug discovery.

Selected figure(s)

Figure 4.
FIGURE 4. Three-dimensional structure of the active site of sortase B modified by AAEK1. A, thienylpropanone adduct to sortase Cys-233 (blue), the product of the reaction of sortase B with AAEK1, is compared with the sortase B structure at 1.6 Å (orange). The adduct occupies the position of three water molecules (orange spheres) and is stacking against Tyr-138. B, the catalytic triad of sortase B (His140, Asp-234, and Cys-233) and Arg-243 are in close proximity to the inhibitor adduct and undergo substantial structural shifts. The SrtB (green) and SrtB-AAEK adduct (blue) are superimposed. The figure was generated using PyMOL^TM. C, thienylpropanone modification of the active site of sortase. Electron density map of the B. anthracis sortase B-AAEK1 adduct demonstrating Cys-233, Tyr-138, and the thienylpropanone. Green, sulfur; red, oxygen. The figure was generated using COOT.

Figure 7.
FIGURE 7. Model for the mechanism of inhibition of sortase by AAEKs. Deprotonation of the carbon is conjectured to occur via a base in the active site of sortase. This generates enolate 13, which may be stabilized in a manner similar to the oxyanion intermediate of sortase during catalysis (e.g. by the guanidinium of a conserved active site arginine). This intermediate eliminates an amine, here dimethylamine, from the -position to generate 14. The electrophilic nature of 14 allows it to serve as an acceptor in a Michael-type conjugate addition by the thiol of the active site cysteine. This resulting enolate might also be stabilized by the guanidinium moiety; subsequent protonation by enzyme or medium would then generate the stable AAEK thioether adduct observed.

The above figures are reprinted by permission from the ASBMB: J Biol Chem (2007, 282, 23129-23139) copyright 2007.

Figures were selected by an automated process.