Allosteric communication in class A β-lactamases occurs via cooperative coupling of loop dynamics
- University College London, United Kingdom
- University of Bristol, United Kingdom
- Case Western Reserve University, United States
- Oklahoma State University, United States
Abstract
Understanding allostery in enzymes and tools to identify it, offer promising alternative strategies to inhibitor development. Through a combination of equilibrium and nonequilibrium molecular dynamics simulations, we identify allosteric effects and communication pathways in two prototypical class A β-lactamases, TEM-1 and KPC-2, which are important determinants of antibiotic resistance. The nonequilibrium simulations reveal pathways of communication operating over distances of 30 Å or more. Propagation of the signal occurs through cooperative coupling of loop dynamics. Notably, 50% or more of clinically relevant amino acid substitutions map onto the identified signal transduction pathways. This suggests that clinically important variation may affect, or be driven by, differences in allosteric behavior, providing a mechanism by which amino acid substitutions may affect the relationship between spectrum of activity, catalytic turnover and potential allosteric behavior in this clinically important enzyme family. Simulations of the type presented here will help in identifying and analyzing such differences.
Data availability
All analysis scripts have been uploaded on figshare with doi 10.6084/m9.figshare.13583384
Article and author information
Author details
Francesco Luigi Gervasio
Adrian J Mulholland
Funding
AstraZeneca (Case Studentship)
- Ioannis Galdadas
National Institute of Allergy and Infectious Diseases (R01AI072219)
- Robert A Bonomo
National Institute of General Medical Sciences (GM105978)
- Pratul K Agarwal
National Institutes of Health (RO1AI063517)
- Robert A Bonomo
- Shozeb Haider
Engineering and Physical Sciences Research Council (EP/M022609/1)
- Ana Sofia F Oliveira
- Adrian J Mulholland
Engineering and Physical Sciences Research Council (EP/N024117/1)
- Ana Sofia F Oliveira
- Adrian J Mulholland
Biotechnology and Biological Sciences Research Council (BB/L01386X/1)
- Ana Sofia F Oliveira
- Adrian J Mulholland
Medical Research Council (MR/T016035/1)
- Catherine L Tooke
- James Spencer
- Adrian J Mulholland
National Institute of Allergy and Infectious Diseases (R01AI100560)
- Robert A Bonomo
National Institute of Allergy and Infectious Diseases (R01AI063517)
- Robert A Bonomo
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Yogesh K Gupta, University of Texas Health Science Center at San Antonio, United States
Version history
- Received: January 14, 2021
- Accepted: March 19, 2021
- Accepted Manuscript published: March 23, 2021 (version 1)
- Accepted Manuscript updated: March 24, 2021 (version 2)
- Version of Record published: April 21, 2021 (version 3)
Copyright
© 2021, Galdadas et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Metrics
-
- 3,814
- views
-
- 495
- downloads
-
- 52
- citations
Views, downloads and citations are aggregated across all versions of this paper published by eLife.
Download links
A two-part list of links to download the article, or parts of the article, in various formats.
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
Allosteric communication in class A β-lactamases occurs via cooperative coupling of loop dynamics
eLife 10:e66567.
https://doi.org/10.7554/eLife.66567
Further reading
-
- Developmental Biology
- Medicine
From a forward mutagenetic screen to discover mutations associated with obesity, we identified mutations in the Spag7 gene linked to metabolic dysfunction in mice. Here, we show that SPAG7 KO mice are born smaller and develop obesity and glucose intolerance in adulthood. This obesity does not stem from hyperphagia, but a decrease in energy expenditure. The KO animals also display reduced exercise tolerance and muscle function due to impaired mitochondrial function. Furthermore, SPAG7-deficiency in developing embryos leads to intrauterine growth restriction, brought on by placental insufficiency, likely due to abnormal development of the placental junctional zone. This insufficiency leads to loss of SPAG7-deficient fetuses in utero and reduced birth weights of those that survive. We hypothesize that a ‘thrifty phenotype’ is ingrained in SPAG7 KO animals during development that leads to adult obesity. Collectively, these results indicate that SPAG7 is essential for embryonic development and energy homeostasis later in life.
-
- Medicine
Background:
The development of obesity-associated comorbidities such as type 2 diabetes (T2D) and hepatic steatosis has been linked to selected microRNAs in individual studies; however, an unbiased genome-wide approach to map T2D induced changes in the miRNAs landscape in human liver samples, and a subsequent robust identification and validation of target genes are still missing.
Methods:
Liver biopsies from age- and gender-matched obese individuals with (n=20) or without (n=20) T2D were used for microRNA microarray analysis. The candidate microRNA and target genes were validated in 85 human liver samples, and subsequently mechanistically characterized in hepatic cells as well as by dietary interventions and hepatic overexpression in mice.
Results:
Here, we present the human hepatic microRNA transcriptome of type 2 diabetes in liver biopsies and use a novel seed prediction tool to robustly identify microRNA target genes, which were then validated in a unique cohort of 85 human livers. Subsequent mouse studies identified a distinct signature of T2D-associated miRNAs, partly conserved in both species. Of those, human-murine miR-182–5 p was the most associated with whole-body glucose homeostasis and hepatic lipid metabolism. Its target gene LRP6 was consistently lower expressed in livers of obese T2D humans and mice as well as under conditions of miR-182–5 p overexpression. Weight loss in obese mice decreased hepatic miR-182–5 p and restored Lrp6 expression and other miR-182–5 p target genes. Hepatic overexpression of miR-182–5 p in mice rapidly decreased LRP6 protein levels and increased liver triglycerides and fasting insulin under obesogenic conditions after only seven days.
Conclusions:
By mapping the hepatic miRNA-transcriptome of type 2 diabetic obese subjects, validating conserved miRNAs in diet-induced mice, and establishing a novel miRNA prediction tool, we provide a robust and unique resource that will pave the way for future studies in the field. As proof of concept, we revealed that the repression of LRP6 by miR-182–5 p, which promotes lipogenesis and impairs glucose homeostasis, provides a novel mechanistic link between T2D and non-alcoholic fatty liver disease, and demonstrate in vivo that miR-182–5 p can serve as a future drug target for the treatment of obesity-driven hepatic steatosis.
Funding:
This work was supported by research funding from the Deutsche Forschungsgemeinschaft (KI 1887/2-1, KI 1887/2-2, KI 1887/3-1 and CRC-TR296), the European Research Council (ERC, CoG Yoyo LepReSens no. 101002247; PTP), the Helmholtz Association (Initiative and Networking Fund International Helmholtz Research School for Diabetes; MB) and the German Center for Diabetes Research (DZD Next Grant 82DZD09D1G).
