The genome of a Bacteroidetes inhabitant of the human gut encodes a structurally distinct enoyl-acyl carrier protein reductase (FabI)

Christopher D. Radka, Matthew W. Frank, Jiangwei Yao, Jayaraman Seetharaman, Darcie J. Miller, Charles O. Rock

Research output: Contribution to journalArticlepeer-review

6 Scopus citations


Enoyl-acyl carrier protein reductase (FabI) catalyzes a rate-controlling step in bacterial fatty-acid synthesis and is a target for antibacterial drug development. A phylogenetic analysis shows that FabIs fall into four divergent clades. Members of clades 1–3 have been structurally and biochemically characterized, but the fourth clade, found in members of phylum Bacteroidetes, is uncharacterized. Here, we identified the unique structure and conformational changes that distinguish clade 4 FabIs. Alistipes finegoldii is a prototypical Bacteroidetes inhabitant of the gut microbiome. We found that A. finegoldii FabI (AfFabI) displays cooperative kinetics and uses NADH as a cofactor, and its crystal structure at 1.72 Å resolution showed that it adopts a Rossmann fold as do other characterized FabIs. It also disclosed a carboxyl-terminal extension that forms a helix–helix interaction that links the protomers as a unique feature of AfFabI. An AfFabI.NADH crystal structure at 1.86 Å resolution revealed that this feature undergoes a large conformational change to participate in covering the NADH-binding pocket and establishing the water channels that connect the active site to the central water well. Progressive deletion of these interactions led to catalytically compromised proteins that fail to bind NADH. This unique conformational change imparted a distinct shape to the AfFabI active site that renders it refractory to a FabI drug that targets clade 1 and 3 pathogens. We conclude that the clade 4 FabI, found in the Bacteroidetes inhabitants of the gut, have several structural features and conformational transitions that distinguish them from other bacterial FabIs.

Original languageEnglish
Pages (from-to)7635-7652
Number of pages18
JournalJournal of Biological Chemistry
Issue number22
StatePublished - May 28 2020

Bibliographical note

Funding Information:
Acknowledgments—We thank Dr. Amanda Nourse (Molecular Interaction Analysis Shared Resource, St. Jude Children’s Research Hospital) for sample analysis by analytical ultracentrifugation and Brett Waddell (Molecular Interaction Analysis Shared Resource, St. Jude Children’s Research Hospital) for NADH-binding experiments by surface plasmon resonance. We thank the St. Jude Structural Biology X-ray Center for crystallography support. The Southeast Regional Collaborative Access Team (SER-CAT) 22-ID beamline at the Advanced Photon Source, Argonne National Laboratory is supported by its member institutions, and equipment grants (S10_RR25528 and S10_RR028976) from the National Institutes of Health. Use of the Advanced Photon Source was supported by the U. S. Department of Energy, Office of Science, Office of Basic Energy Sciences, under Contract No. W-31–109-Eng-38. The NYX (19-ID) and AMX (17-ID-1) beamlines of the National Synchrotron Light Source II is a U.S. Department of Energy (DOE) Office of Science User Facility operated for the DOE Office of Science by Brookhaven National Laboratory under Contract DE-SC0012704. NYX is supported by the New York Structural Biology Center (NYSBC). AMX is part of the Center for BioMolecular Structure that is supported by the National Institute of General Medical Sciences (P30 GM133893), and by the DOE Office of Biological and Environmental Research (KP 1605010).

Funding Information:
This work was supported by NIGMS, National Institutes of Health Grant GM034496 (to C. O. R.), Cancer Center Support Grant CA21765, and the American Lebanese Syrian Associated Charities. The authors declare that they have no conflicts of interest with the contents of this article. The con-tent is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

Publisher Copyright:
© 2020 Radka et al. Published under exclusive license by The American Society for Biochemistry and Molecular Biology, Inc.

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology


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