Mutation S115T in IMP-Type Metallo-β-Lactamases Compensates for Decreased Expression Levels Caused by Mutation S119G.


Abstract

(1) Background: Metallo-β-lactamases (MBLs) have raised concerns due to their ability to inactivate carbapenems and newer generation cephalosporins and the absence of clinically available MBL inhibitors. Their genes are often transferred horizontally, and the number of MBL variants has grown exponentially, with many newer variants showing enhanced enzyme activity or stability. In this study, we investigated a closely related group of variants from the IMP family that all contain the combination of mutations S115T and S119G relative to IMP-1. (2) Methods: The effects of each individual mutation and their combination in the IMP-1 sequence background in comparison to IMP-1 were investigated. Their ability to confer resistance and their in-cell expression levels were determined. All enzymes were purified, and their secondary structure and thermal stability were determined with circular dichroism. Their Zn(II) content and kinetic constants with a panel of β-lactam antibiotics were determined. (3) Results: All four enzymes were viable and conferred resistance to all antibiotics tested except aztreonam. However, the single-mutant enzymes were slightly deficient, IMP-1S115T due to decreased enzyme activity and IMP-1-S119G due to decreased thermal stability and expression, while the double mutant did not show these defects. (4) Conclusions: These observations suggest that S119G was acquired due to its increased enzyme activity and S115T to suppress the thermal stability and expression defect introduced by S119G.

Submission Details

ID: 3QSSDqeA4

Submitter: Paulie Dang

Submission Date: Jan. 14, 2020, 11:51 a.m.

Version: 1

Publication Details
Zhang CJ;Faheem M;Dang P;Morris MN;Kumar P;Oelschlaeger P,Biomolecules (2019) Mutation S115T in IMP-Type Metallo-β-Lactamases Compensates for Decreased Expression Levels Caused by Mutation S119G. PMID:31718049
Additional Information

Explanation of amino acid numbering: Residues 76 and 80 that were mutated in this study are called residues 115 and 119, respectively, according to the standard numbering scheme (PMID 15215079). ND indicates not detectable.

Structure view and single mutant data analysis

Study data

No weblogo for data of varying length.
Colors: D E R H K S T N Q A V I L M F Y W C G P
 

Data Distribution

Studies with similar sequences (approximate matches)

Correlation with other assays (exact sequence matches)


Relevant PDB Entries

Structure ID Release Date Resolution Structure Title
6JED 2019-08-07 1.57 Crystal structure of IMP-1 metallo-beta-lactamase in a complex with MCR
6LBL 2020-02-19 1.68 Crystal structure of IMP-1 metallo-beta-lactamase in complex with NO9 inhibitor
5Y5B 2018-08-08 1.7 Crystal Structure Of IMP-1 Metallo-beta-lactamase
4C1G 2014-08-27 1.71 Crystal structure of the metallo-beta-lactamase IMP-1 with D-captopril
5EV6 2016-06-01 1.98 Crystal structure of the native, di-zinc metallo-beta-lactamase IMP-1
1DD6 2000-11-08 2.0 IMP-1 METALLO BETA-LACTAMASE FROM PSEUDOMONAS AERUGINOSA IN COMPLEX WITH A MERCAPTOCARBOXYLATE INHIBITOR
5HH4 2017-01-18 2.0 Crystal structure of metallo-beta-lactamase IMP-1 in complex with a phosphonate-based inhibitor
1WUO 2005-03-29 2.01 Crystal structure of metallo-beta-lactamase IMP-1 mutant (D81A)
6JKA 2020-03-25 2.01 Crystal structure of metallo-beta-lactamse, IMP-1, in complex with a thiazole-bearing inhibitor
4C1F 2014-08-27 2.01 Crystal structure of the metallo-beta-lactamase IMP-1 with L-captopril
5EV8 2016-06-01 2.3 Crystal structure of the metallo-beta-lactamase IMP-1 in complex with the bisthiazolidine inhibitor D-CS319
5EWA 2016-06-01 2.3 Crystal structure of the metallo-beta-lactamase IMP-1 in complex with the bisthiazolidine inhibitor L-VC26
2DOO 2006-11-28 2.43 The structure of IMP-1 complexed with the detecting reagent (DansylC4SH) by a fluorescent probe
1VGN 2005-06-21 2.63 Structure-based design of the irreversible inhibitors to metallo--lactamase (IMP-1)
1WUP 2005-03-29 3.0 Crystal structure of metallo-beta-lactamase IMP-1 mutant (D81E)

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
100.0 Metallo-beta-lactamase type 2 P52699 BLAB_SERMA