A Kinetic Study of the Replacement by Site Saturation Mutagenesis of Residue 119 in NDM-1 Metallo-β-Lactamase.
Abstract
New Delhi metallo-β-lactamase 1 (NDM-1) is a subclass B1 metallo-β-lactamase that exhibits a broad spectrum of activity against β-lactam antibiotics. Here we report the kinetic study of 6 Q119X variants obtained by site-directed mutagenesis of NDM-1. All Q119X variants were able to hydrolyze carbapenems, penicillins and first-, second-, third-, and fourth-generation cephalosporins very efficiently. In particular, Q119E, Q119Y, Q119V, and Q119K mutants showed improvements in kcat/Km values for penicillins, compared with NDM-1. The catalytic efficiencies of the Q119K variant for benzylpenicillin and carbenicillin were about 65- and 70-fold higher, respectively, than those of NDM-1. The Q119K and Q119Y enzymes had kcat/Km values for ceftazidime about 25- and 89-fold higher, respectively, than that of NDM-1.
Submission Details
ID: CWvASvaZ3
Submitter: Shu-Ching Ou
Submission Date: March 20, 2019, 3:27 p.m.
Version: 1
Publication Details
Marcoccia F;Mercuri PS;Galleni M;Celenza G;Amicosante G;Perilli M,Antimicrob Agents Chemother (2018) A Kinetic Study of the Replacement by Site Saturation Mutagenesis of Residue 119 in NDM-1 Metallo-β-Lactamase. PMID:29784851Additional Information
Study Summary
| Number of data points | 299 |
| Proteins | Metallo-beta-lactamase type 2 |
| Unique complexes | 117 |
| Assays/Quantities/Protocols | Experimental Assay: MIC ; Experimental Assay: kcat ; Experimental Assay: Km ; Experimental Assay: kcat/Km |
| Libraries | Variants for NDM-1_Substrate |
Structure view and single mutant data analysis
Study data
| 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 |
|---|---|---|---|
| 5ZJ1 | 2018-03-18T00:00:00+0000 | 0.96 | Crystal structure of NDM-1 in complex with D-captopril derivative CYT-14 |
| 6C6I | 2018-01-18T00:00:00+0000 | 1.65 | Crystal structure of a chimeric NDM-1 metallo-beta-lactamase harboring the IMP-1 L3 loop |
| 5ZJ2 | 2018-03-18T00:00:00+0000 | 1.1 | Crystal structure of NDM-1 in complex with D-captopril |
| 3PG4 | 2010-10-29T00:00:00+0000 | 2.0 | The crystal structure of New Delhi Metallo-beta lactamase (NDM-1) |
| 5ZJ7 | 2018-03-19T00:00:00+0000 | 0.96 | Crystal structure of NDM-1 in complex with D-captopril derivative CY22 |
| 5ZJ8 | 2018-03-19T00:00:00+0000 | 0.96 | Crystal structure of NDM-1 in complex with D-captopril derivative CY32 |
| 5ZGP | 2018-03-10T00:00:00+0000 | 1.15 | Crystal structure of NDM-1 at pH6.2 (Bis-Tris) in complex with hydrolyzed ampicillin |
| 5ZH1 | 2018-03-10T00:00:00+0000 | 1.05 | Crystal structure of NDM-1 at pH7.5 (Imidazole) with 2 molecules per asymmetric unit |
| 6LJ7 | 2019-12-13T00:00:00+0000 | 1.16 | Crystal structure of NDM-1 in complex with D-captopril derivative wss05010 |
| 3SBL | 2011-06-05T00:00:00+0000 | 2.31 | Crystal Structure of New Delhi Metal-beta-lactamase-1 from Klebsiella pneumoniae |
Relevant UniProtKB Entries
| Percent Identity | Matching Chains | Protein | Accession | Entry Name |
|---|---|---|---|---|
| 100.0 | Metallo-beta-lactamase type 2 | C7C422 | BLAN1_KLEPN |