Hydroxyl groups in the (beta)beta sandwich of metallo-beta-lactamases favor enzyme activity: a computational protein design study.


Abstract

Metallo-beta-lactamases challenge antimicrobial therapies by their ability to hydrolyze and inactivate a broad spectrum of beta-lactam antibiotics. The potential of these enzymes to acquire enhanced catalytic efficiency through mutation is of great concern. Here, we explore the potential of computational protein design to predict mutants of the imipenemase IMP-1 that modulate the catalytic efficiency of the enzyme against a range of substrates. Focusing on the four amino acid positions 69, 121, 218, and 262, we carried out a number of design calculations. Two mutant enzymes were predicted: the single mutant S262A and the double mutant F218Y-S262A. Compared to IMP-1, the single mutant (S262A) results in the loss of a hydroxyl group and the double mutant (F218Y-S262A) results in a hydroxyl transfer from position 262 to position 218. The presence of both hydroxyl groups at positions 218 and 262 was tested by examining the mutant F218Y. Kinetic constants of IMP-1, the two computationally designed mutants (S262A and F218Y-S262A), and the hydroxyl addition mutant (F218Y) were determined with seven substrates. Catalytic efficiencies are highest for the enzyme with both hydroxyl groups (F218Y) and lowest for the enzyme lacking both hydroxyl groups (S262A). The catalytic efficiencies of the two enzymes with one hydroxyl group each are intermediate, with the F218Y-S262A double mutant exhibiting enhanced hydrolysis of nitrocefin, cephalothin, and cefotaxime relative to IMP-1.

Submission Details

ID: H5223AJS

Submitter: Peter Oelschlaeger

Submission Date: Jan. 9, 2019, 9:23 a.m.

Version: 1

Publication Details
Oelschlaeger P;Mayo SL,J Mol Biol (2005) Hydroxyl groups in the (beta)beta sandwich of metallo-beta-lactamases favor enzyme activity: a computational protein design study. PMID:15946681
Additional Information

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
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
5HH4 2017-01-18 2.0 Crystal structure of metallo-beta-lactamase IMP-1 in complex with a phosphonate-based inhibitor
1DD6 2000-11-08 2.0 IMP-1 METALLO BETA-LACTAMASE FROM PSEUDOMONAS AERUGINOSA IN COMPLEX WITH A MERCAPTOCARBOXYLATE INHIBITOR
1WUO 2005-03-29 2.01 Crystal structure of metallo-beta-lactamase IMP-1 mutant (D81A)
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
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