[john innes centre, norwich]

Ghilarov Lab

We study bacterial molecular machines to make better antibiotics

We stand with Ukraine. Aggressor must be stopped. Нет войне!

Based at the John Innes Centre, Norwich, the world-renowned place for antibiotic research. We apply cryo-electron microscopy to discover how bacteria make, transport and acquire resistance to antibiotics to deliver better healthcare for the future world. Promoting fair, kind and inclusive academic culture.

News

Research

Our main research focus is on the biosynthesis and activity of peptide antibiotics, mostly ribosomally synthesised (RiPPs). We are structural biologists and we use cryo-electron microscopy (cryo-EM) to dissect the catalytic mechanisms of biosynthetic enzymes: these can be subsequently used to create new to nature bioactive compounds. We are also pursuing mode of action of known antibiotics, with a particular focus on bacterial DNA topoisomerases inhibition. We have a range of other research projects & collaborations in the broader field of molecular microbiology: anti-phage defense mechanisms, peptide transporters, antibiotic resistance proteins.

Microcin B17 biosynthesis complex McbBCD [https://doi.org/10.1016/j.molcel.2018.11.032]

BIOSYNTHESIS OF RIBOSOMALLY SYNTHESIZED ANTIBIOTICS

A large group of bacterial antibiotics are produced by enzymatic transformations of ribosomally-synthesized precursors. Such compounds are termed "RiPPs" for ribosomally-synthesized post-translationally modified peptides.

YcaO enzymes are responsible for peptide backbone modifications in RiPPs: conversion of amino acids into heterocycles and thioamidation (replacement of peptide bond oxygen by sulfur). Peptide bonds are generally very stable and difficult to chemically modify, but YcaOs do so easily. YcaOs are capable of modifying virtually any substrate peptide bearing a correct recognition sequence, making them highly valuable for industry.

We work with microcin B17 and klebsazolicin RiPP systems to unravel the mechanistic details of YcaO-catalysed reactions.

ANTIBIOTIC TRANSPORT

To make an effective drug, we need to know how to deliver it to the target cell.
Bacterial outer and inner membrane transporters are key to the sensitivity to antibacterial peptides. In collaboration with the groups of Konstantinos Beis, Christos Pliotas and Jonathan Heddle we study the molecular mechanisms of antibiotic transport and determinants for the compound selectivity. We use lipid membrane mimetics (nanodiscs) and cryo-electron microscopy to look at the transporters in the conditions most close to native.

Escherichia coli inner membrane transporter SbmA [https://doi.org/10.1126/sciadv.abj5363]

DNA GYRASE & NEW ANTIBIOTICS

Mode of action of microcin B17 and many other peptide antibiotics targeting DNA gyrase is not known. By determining cryo-EM structures of DNA gyrase in complex with these compounds, we are preparing grounds for the development of new antimicrobials targeting different enzyme pockets and thus avoiding existing resistance.

E.coli DNA gyrase cryo-EM map determined in the lab

Srtucture of K. pneumoniae QnrB1 determined by cryo-EM in the lab

DNA GYRASE & FLUOROQUINOLONE RESISTANCE

DNA gyrase inhibitors fluoroquinolones are one of the most prescribed antibiotics. Bacterial genomes encode special proteins to endow gyrase with a degree of resistance to these drugs. We are applying cryo-EM to study molecular mechanisms of this process.

People

Yinghong Gu
Postdoctoral Scientist
Yinghong comes from Chongqing, a municipality in southwest China. He has deep expertise in membrane protein structural biology. He uses cryo-EM methods to reveal the modification mechanisms of a class of natural peptides, RiPPs, and develop them into novel antimicrobial agents.
Dmitry Ghilarov
Group Leader, Wellcome Trust Sir Henry Dale Fellow
Dmitry did his PhD in Russia and worked as visiting scientist at the John Innes Centre before moving to Poland and now returning to the UK to set up his own lab. Throughout his career, Dmitry studied RiPPs microcin B17 and klebsazolicin, their biosynthesis and target of microcin B17 - DNA gyrase.
Leela Ghimire
Research Assistant
After doing her BSc in the Philippines and MSc in Leeds, UK Leela joined us to investigate a new class of topoisomerase inhibitors through biochemical assays and cryo-EM
Myfanwy Adams
Postdoctoral Scientist
Myfanwy did her structural biology PhD in Cornell and joined us to jump into cryo-EM field. She is working on bacterial anti-phage defense systems and RiPPs biosynthesis.
Maria Mihaylenko
Research Intern
Maria finished her BSc in the University of East Anglia and works as an intern to help with protein purification for marine RiPP systems

Funding

Publications

[2023] Molecular mechanism of topoisomerase poisoning by the peptide antibiotic albicidin

In collaboration with Roderich Sussmuth from TU Berlin and Jonathan Heddle from UJ Krakow we used cryo-EM to determine mode of action of a potent antibiotic albicidin. Albicidin inhibits topoisomerase by a novel mechanism,: one part of the molecule slides in between gyrase main dimer-dimer interface, whilst another part intercalates into DNA

[2021] Molecular mechanism of SbmA, a promiscuous transporter exploited by antimicrobial peptides

In collaboration with Dr. Konstantinos Beis from Imperial College London we solved cryo-EM structure of an E.coli inner membrane transporter SbmA responsible for the entry of Microcin B17 and other antibacterial peptides inside the cells.

[2021] Pentapeptide repeat protein QnrB1 requires ATP hydrolysis to rejuvenate poisoned gyrase complexes

Here we investigated the mechanism of action of fluoroquinolone resistance factor QnrB1 which binds to the DNA gyrase causing release of the drug [doi: 10.1093/nar/gkaa1266]

[2019] Architecture of Microcin B17 Synthetase: An Octameric Protein Complex Converting a Ribosomally Synthesized Peptide into a DNA Gyrase Poison

Molecular Cell Feb 2019 cover. Here we determined the crystal structure of the microcin B17 synthetase McbBCD, which offered the first glimpse of a complete RiPP synthetic machinery [https://doi.org/10.1016/j.molcel.2018.11.032]

[2018] Biosynthesis of Translation Inhibitor Klebsazolicin Proceeds through Heterocyclization and N-Terminal Amidine Formation Catalyzed by a Single YcaO Enzyme

We reconstituted biosynthesis of the novel RiPP klebsazolicin discovered in Konstantin Severinov Lab and showed that a single enzyme catalyses both heterocyclisation and macroamidine formation in klebsazolicin [https://doi.org/10.1021/jacs.8b02277]

[2017] The Origins of Specificity in the Microcin-Processing Protease TldD/E

Here we determined the structure of and characterise a highly conserved bacterial protease TldD/E responsible for the maturation of several ribosomally synthesized antibiotics [https://doi.org/10.1016/j.str.2017.08.006]

Join Us!

We are looking forward to meet inspired people who are not afraid to address global challenges.

PhD students are normally expected to apply for one of Norwich Research Park DTPs or JIC Rotation Programme

We will welcome researchers wishing to apply for Postdoctoral Fellowships such as EMBO, BBSRC Discovery or Marie Curie

Our Colleagues: Laboratory of Biochemistry and Bionanoscience - http://www.heddlelab.org

Contact Us:
Dmitry.Ghilarov@jic.ac.uk

Address
John Innes Centre, Norwich Research Park
Norwich NR4 7UH United Kingdom