Elsevier

Drug Resistance Updates

Volume 40, September 2018, Pages 1-12
Drug Resistance Updates

A close look onto structural models and primary ligands of metallo-β-lactamases

https://doi.org/10.1016/j.drup.2018.08.001Get rights and content

Abstract

β-Lactamases are hydrolytic enzymes capable of opening the β-lactam ring of antibiotics such as penicillin, thus endowing the bacteria that produce them with antibiotic resistance. Of particular medical concern are metallo-β-lactamases (MBLs), with an active site built around coordinated Zn cations. MBLs are pan-reactive enzymes that can break down almost all classes of β-lactams, including such last-resort antibiotics as carbapenems. They are not only broad-spectrum-reactive but are often plasmid-borne (e.g., the New Delhi enzyme, NDM), and can spread horizontally even among unrelated bacteria. Acquired MBLs are encoded by mobile genetic elements, which often include other resistance genes, making the microbiological situation particularly alarming. There is an urgent need to develop MBL inhibitors in order to rescue our antibiotic armory. A number of such efforts have been undertaken, most notably using the 3D structures of various MBLs as drug-design targets. Structure-guided drug discovery depends on the quality of the structures that are collected in the Protein Data Bank (PDB) and on the consistency of the information in dedicated β-lactamase databases. We conducted a careful review of the crystal structures of class B β-lactamases, concluding that the quality of these structures varies widely, especially in the regions where small molecules interact with the macromolecules. In a number of examples the interpretation of the bound ligands (e.g., inhibitors, substrate/product analogs) is doubtful or even incorrect, and it appears that in some cases the modeling of ligands was not supported by electron density. For ten MBL structures, alternative interpretations of the original diffraction data could be proposed and the new models have been deposited in the PDB. In four cases, these models, prepared jointly with the authors of the original depositions, superseded the previous deposits. This review emphasizes the importance of critical assessment of structural models describing key drug design targets at the level of the raw experimental data. Since the structures reviewed here are the basis for ongoing design of new MBL inhibitors, it is important to identify and correct the problems with ambiguous crystallographic interpretations, thus enhancing reproducibility in this highly medically relevant area.

Section snippets

Introduction – reproducibility crisis in biomedical research

A number of recent reports have brought to the attention of the scientific community the uncomfortable fact that a noticeable fraction of biomedical research cannot be reproduced (Minor et al., 2016; Prinz et al., 2011). This is an alarming trend, and even more alarming outlook, calling for immediate action. Not only are large resources of time, manpower, and money (estimated at $28 billion a year in the US alone (Freedman et al., 2015)) invested in potentially useless endeavors, but also false

Model validation methods

No new experimental data were collected for this work and all analyses were based on data obtained by others and deposited in the PDB and/or https://proteindiffraction.org/. We reviewed over 150 deposited crystal structures of metallo-β-lactamases and we used the BLDB database (Naas et al., 2017) to identify the relevant structures of B1, B2, and B3 β-lactamases. Only the entries containing experimental diffraction data were subjected to our detailed review.

When available, electron density maps

NDM-1 with tiopronin (PDB ID 5a5z)

A 2.6 Å structure of NDM-1, supposedly with the ligand tiopronin bound at the active site, was determined as part of a study entitled “Approved drugs containing thiols as inhibitors of metallo-β-lactamases: Strategy to combat multidrug-resistant bacteria” (Klingler et al., 2015). When this work was initially published, there was no accompanying PDB deposit. After our correspondence with the author and, subsequently, with the journal editor, the authors submitted the model coordinates and the

The ripple effect of sub-optimal and incorrect structural models

Crystallographic structural data, once deposited in the PDB and reported in the literature, tend to be treated by the non-structural community with full trust and their correctness is rarely questioned. The “ripple effect” of structural models (good and bad!) is very significant, given that many other fields of science use these models as the foundation for their research. Protein crystallography gives scientists an unprecedented power to investigate life at sub-microscopic levels, by revealing

Acknowledgments

We wish to thank Dr. Bernhard Rupp for valuable comments on one of the cases (5a5z). We also acknowledge, with thanks, the cooperation and joint authorship of the revised PDB deposits on the part of some of the original authors, especially Dr. Andrzej Joachimiak, Dr. Youngchang Kim, Dr. Natalie Strynadka, Dr. Dustin King, Dr. Alejandro Vila, Dr. Javier Gonzalez, Dr. Stefano Mangani, Dr. Isabel Garcia-Saez, Dr. Jean-Marie Frere, and Dr. Otto Dideberg. The work of MJ and JR was supported by the

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      Citation Excerpt :

      The di-zinc B1 subclass MBLs (e.g., NDM, VIM, and IMP types) are of widespread clinical significance10–12. They are characterized by a broad substrate specificity and able to hydrolyze almost all β-lactam antibiotics including carbapenems13–14. MBL inhibitors targeting these di-zinc MBLs have been widely developed in recent years, mainly acting through active site metal chelating and metal deprivation15–17.

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    Dedication: As a birthday tribute, this work is dedicated to Dr. Zbigniew Dauter, an untiring advocate of the highest quality in structural research.

    1

    Equal contribution.

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