PDBsum entry 6j2b

Hydrolase/inhibitor

PDB id: 6j2b

Contents

Protein chains

260 a.a.

Ligands

TSL ×2

GOL ×2

Waters ×451

PDB id:

6j2b

Name:

Hydrolase/inhibitor

Title:

Ctx-m-64 beta-lactamase s130t sulbactam complex

Structure:

Beta-lactamase. Chain: a, b. Fragment: unp residues 29-291. Engineered: yes. Mutation: yes

Source:

Escherichia coli. Organism_taxid: 562. Gene: blactx-m-64. Expressed in: escherichia coli 'bl21-gold(de3)plyss ag'. Expression_system_taxid: 866768

Resolution:

1.44Å R-factor: 0.154 R-free: 0.187

Authors:

Q.Cheng,S.Chen

Key ref:

Q.Cheng et al. (2020). Structural Insight into the Mechanism of Inhibitor Resistance in CTX-M-199, a CTX-M-64 Variant Carrying the S¹³⁰T Substitution. ACS Infect Dis, 6, 577-587. PubMed id: 31709791 DOI: 10.1021/acsinfecdis.9b00345

Date:

31-Dec-18 Release date: 30-Oct-19

Protein chains

C8CP57  (C8CP57_ECOLX) -  Beta-lactamase from Escherichia coli

Seq: 291 a.a.

Struc: 260 a.a.*

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

Enzyme reactions

• Enzyme class: E.C.3.5.2.6 - beta-lactamase.
• Pathway: Penicillin Biosynthesis and Metabolism
• Reaction: a beta-lactam + H2O = a substituted beta-amino acid
• Cofactor: Zn(2+)

DOI no: 10.1021/acsinfecdis.9b00345

ACS Infect Dis 6:577-587 (2020)

PubMed id: 31709791

Structural Insight into the Mechanism of Inhibitor Resistance in CTX-M-199, a CTX-M-64 Variant Carrying the S¹³⁰T Substitution.

Q.Cheng, C.Xu, J.Chai, R.Zhang, E.Wai Chi Chan, S.Chen.

ABSTRACT

The smart design of β-lactamase inhibitors allowed us to combat extended-spectrum β-lactamase (ESBL)-producing organisms for many years without developing resistance to these inhibitors. However, novel resistant variants have emerged recently, and notable examples are the CTX-M-190 and CTX-M-199 variants, which carried a S¹³⁰T amino acid substitution and exhibited resistance to inhibitors such as sulbactam and tazobactam. Using mass spectrometric and crystallographic approaches, this study depicted the mechanisms of inhibitor resistance. Our data showed that CTX-M-64 (S¹³⁰T) did not cause any conformational change or exert any effect on its ability to hydrolyze β-lactam substrates. However, binding of sulbactam, but not clavulanic acid, to the active site of CTX-M-64 (S¹³⁰T) led to the conformational changes in such active site, which comprised the key residues involved in substrate catalysis, namely, Thr¹³⁰, Lys⁷³, Lys²³⁴, Asn¹⁰⁴, and Asn¹³². This conformational change weakened the binding of the sulbactam trans-enamine intermediate (TSL) to the active site and rendered the formation of the inhibitor-enzyme complex, which features a covalent acrylic acid (AKR)-T¹³⁰ bond, inefficient, thereby resulting in inhibitor resistance in CTX-M-64 (S¹³⁰T). Understanding the mechanisms of inhibitor resistance provided structural insight for the future development of new inhibitors against inhibitor-resistant β-lactamases.