Structure of Geobacter OmcZ filaments suggests extracellular cytochrome polymers evolved independently multiple times

  1. Fengbin Wang
  2. Chi Ho Chan
  3. Victor Suciu
  4. Khawla Mustafa
  5. Madeline Ammend
  6. Dong Si
  7. Allon I Hochbaum
  8. Edward H Egelman
  9. Daniel R Bond

  • Curated by eLife

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    Evaluation Summary:

    This manuscript reports the CryoEM structure of OmcZ cytochrome nanowires of Geobacter sulfurreducens, the third cytochrome nanowire of Geobacter to be structurally resolved. OmcZ differs structurally from these previously determined nanowire structures, showing a different heme chain configuration. Based on these and other differences the authors speculate about the evolutionary origin of these nanowires and the mechanism of long-range electron transport. This manuscript is an important contribution to the field of electron transfer and will be of interest to everyone working in electron transfer and filament formation and interested in their evolution.

    (This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. Reviewer #1 agreed to share their name with the authors.)

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Abstract

While early genetic and low-resolution structural observations suggested that extracellular conductive filaments on metal-reducing organisms such as Geobacter were composed of type IV pili, it has now been established that bacterial c -type cytochromes can polymerize to form extracellular filaments capable of long-range electron transport. Atomic structures exist for two such cytochrome filaments, formed from the hexaheme cytochrome OmcS and the tetraheme cytochrome OmcE. Due to the highly conserved heme packing within the central OmcS and OmcE cores, and shared pattern of heme coordination between subunits, it has been suggested that these polymers have a common origin. We have now used cryo-electron microscopy (cryo-EM) to determine the structure of a third extracellular filament, formed from the Geobacter sulfurreducens octaheme cytochrome, OmcZ. In contrast to the linear heme chains in OmcS and OmcE from the same organism, the packing of hemes, heme:heme angles, and between-subunit heme coordination is quite different in OmcZ. A branched heme arrangement within OmcZ leads to a highly surface exposed heme in every subunit, which may account for the formation of conductive biofilm networks, and explain the higher measured conductivity of OmcZ filaments. This new structural evidence suggests that conductive cytochrome polymers arose independently on more than one occasion from different ancestral multiheme proteins.

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  1. Version published to 10.7554/elife.81551 on eLife
  2. eLife

    Evaluation Summary:

    This manuscript reports the CryoEM structure of OmcZ cytochrome nanowires of Geobacter sulfurreducens, the third cytochrome nanowire of Geobacter to be structurally resolved. OmcZ differs structurally from these previously determined nanowire structures, showing a different heme chain configuration. Based on these and other differences the authors speculate about the evolutionary origin of these nanowires and the mechanism of long-range electron transport. This manuscript is an important contribution to the field of electron transfer and will be of interest to everyone working in electron transfer and filament formation and interested in their evolution.

    (This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. Reviewer #1 agreed to share their name with the authors.)

  3. eLife

    Reviewer #1 (Public Review):

    The authors have determined the structure of OmcZ cytochrome nanowires of Geobacter sulfurreducens by Cryo-EM.

    OmcZ represents the third cytochrome nanowire of Geobacter to be structurally resolved. The structure reveals an octaheme cytochrome which oligomerizes to form an extended filament which scaffolds a continuous chain of hemes which serves to support long-range electron transport to terminal electron acceptors.

    Previously identified nanowires which have been structurally resolved consisted of oligomers of OmcS and OmcE which, although lacking significant sequence identity, shared a common heme arrangement along the filament/nanowire.

    OmcZ differs structurally from OmcS and OmcE, possessing a notably different heme chain configuration. OmcZ also differs from OmcS/OmcE in the nature of the interactions at the interface between subunits. Whilst in OmcS/OmcE a terminal heme is ligated by a histidine from the adjacent subunit of the wire, in OmcZ the terminal heme is ligated by a histidine within the same subunit, highlighting yet another difference between OmcZ and OmcS/E.

    Based upon these observations, the authors suggest that OmcS and OmcE evolved from a common ancestor and that OmcZ evolved independently of OmcS/E. This is significant as it not only reveals the diversity of cytochrome nanowires which support long range electron transfer in Geobacter but also demonstrates that this mechanism of EET has potentially evolved multiple times and is likely to be exploited by other environmental microbes which utilize extracellular electron transport to support respiration.

    Manuscript Strengths:
    The manuscript presents a solid detailed structural analysis of OmcZ providing new insight into the diverse range of electron transfer pathways utilized by Geobacter. By comparing OmcZ with other cytochrome nanowires of Geobacter (OmcS/OmcE) and with other electron transfer proteins such as the MtrABC complex, additional insight is gained into potential electron transfer properties of this cytochrome nanowire.

    Manuscript Weaknesses:
    The manuscript compares previous characterisations of OmcZ filaments by X-ray scattering/IR nanospectroscopy prepared at pH 2 and pH 7 which indicated a higher percentage of alpha-helices and beta-sheets than what was observed by Cryo-EM from filaments prepared at pH 10.5 (this study). Due to the differences observed, it is suggested these previously utilized techniques are unreliable. Although there is a substantial difference in the proportion of beta-sheet that is observed/indicated between different methods, without a direct comparison available at the same pH it is perhaps not possible to attribute differences to the techniques alone.

    Manuscript Impact:
    Through this work, the authors have made a significant contribution to the knowledge surrounding the electron transfer processes of Geobacter. Based on the structure obtained, they have sought to rationalise observed phenotypes associated with the different cytochrome nanowires and intriguingly propose how OmcZ may allow for more conductive biofilms through the formation of meshes of OmcZ filaments capable of exchanging electrons at solvent exposed hemes.

    This manuscript will be of interest to scientists working across a range of disciplines including environmental microbiologists studying microbially driven redox processes in the subsurface, biochemists studying electron transfer proteins/pathways and in particular those working on extracellular electron transfer, and biotechnologists seeking to exploit bacterial electron transfer processes for biotechnological applications.

  4. eLife

    Reviewer #2 (Public Review):

    Wang and co-workers used cryo-electron microscopy to determine a structural model for an extracellular filament from the bacterium Geobacter sulfurreducens. The filament is formed by the polymeric assemble of an octaheme cytochrome, designated OmcZ, and the authors claim that it presents unprecedented structural features compared to other two filaments. The main differences include the heme geometries in each subunit, as well as the structural assemble of the subunits in the polymer. Another important observation encompasses the high solvent exposure of a branched heme that might be important for the higher electron conductivity of OmcZ filament compared the OmcS and OmcE ones.
    Although the paper does have strengths as described above, the weaknesses are that many of the strengths are not directly demonstrated. Strong evidence to support the hypotheses should be included or at least to better consubstantiate them. In particular (a) distinguish between the structural experimental data and modulation/manual curation; (b) extend the discussion of solvent exposition of the cofactors ; (c) clarify the protein's glycosylation sites; (d) critically discuss the experimental conditions in which the structure of the OmcZ filament was observed versus the physiological bacterial growth conditions; (e) discuss the structural determinants for the protein-protein interface stabilization in the filament.

  5. Version published to 10.1101/2022.07.17.500359v1 on bioRxiv