Structural basis for different membrane-binding properties of E. coli anaerobic and human mitochondrial β-oxidation trifunctional enzymes

Sah‐Teli, Shiv K.; Pinkas, Matyáš; Hynönen, Mikko J.; Butcher, Sarah J.; Wierenga, Rik K.; Nováček, Jiří; Radha, Venkatesan

doi:10.1016/j.str.2023.04.011

Cited by 4 publications

(6 citation statements)

References 56 publications

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“…The structural superimposition of the model with the corresponding PDB entry visually confirms the accuracy of the model. The next one is a 10.3 Å map from anaerobic fatty acid beta oxidation trifunctional enzyme (anEcTFE) octameric complex (EMD-16134) 53 . The complex has eight chains, which is a dimer of a tetramer, shown as left and right volumes in the map in the figure.…”

Section: Structure Modeling On Cryo-em Maps At a Lower Resolution Rangementioning

confidence: 99%

DiffModeler: Large Macromolecular Structure Modeling in Low-Resolution Cryo-EM Maps Using Diffusion Model

Wang,

Zhu,

Terashi

et al. 2024

Preprint

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Cryogenic electron microscopy (cryo-EM) has now been widely used for determining multi-chain protein complexes. However, modeling a complex structure is challenging particularly when the map resolution is low, typically in the intermediate resolution range of 5 to 10 Å. Within this resolution range, even accurate structure fitting is difficult, let alone de novo modeling. To address this challenge, here we present DiffModeler, a fully automated method for modeling protein complex structures. DiffModeler employs a diffusion model for backbone tracing and integrates AlphaFold2-predicted single-chain structures for structure fitting. Extensive testing on cryo-EM maps at intermediate resolutions demonstrates the exceptional accuracy of DiffModeler in structure modeling, achieving an average TM-Score of 0.92, surpassing existing methodologies significantly. Notably, DiffModeler successfully modeled a protein complex composed of 47 chains and 13,462 residues, achieving a high TM-Score of 0.94. Further benchmarking at low resolutions (10-20 Å) confirms its versatility, demonstrating plausible performances. Moreover, when coupled with CryoREAD, DiffModeler excels in constructing protein-DNA/RNA complex structures for near-atomic resolution maps (0-5 Å), showcasing state-of-the-art performance with average TM-Scores of 0.88 and 0.91 across two datasets.

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Section: Structure Modeling On Cryo-em Maps At a Lower Resolution Rangementioning

confidence: 99%

DiffModeler: Large Macromolecular Structure Modeling in Low-Resolution Cryo-EM Maps Using Diffusion Model

Wang,

Zhu,

Terashi

et al. 2024

Preprint

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“…Mtb has several copies of genes coding for the enzymes of the β-oxidation cycle; however, only a single trifunctional enzyme (MtTFE) that catalyzes the last three out of four reactions of the β-oxidation cycle (Fig. 1), is present in Mtb, in contrast to other bacteria such as Escherichia coli (E. coli) which has two TFEs (Sah-Teli et al, 2023. MtTFE has been characterized to be an α2β2 heterotetrameric multifunctional enzyme complex (Fig.…”

Section: Introductionmentioning

confidence: 99%

“…Each of the -oxidation TFEs form α2β2 heterotetramers (M=240kDa) in which the β2 dimer forms the core of the tetramer, however, their quaternary assemblies differ, due to different modes of assembly of the α chains onto the β2 dimer (Ishikawa et al, 2004;Liang et al, 2018;Sah-Teli et al, 2023Venkatesan & Wierenga, 2013;Xia et al, 2019). MtTFE, EcTFE and PfTFE are soluble enzymes whereas HsTFE and anEcTFE are membrane associated.…”

Section: Introductionmentioning

confidence: 99%

“…MtTFE, EcTFE and PfTFE are soluble enzymes whereas HsTFE and anEcTFE are membrane associated. The different quaternary assemblies of the four TFE subfamilies, the electrostatic surface features between the active sites, as well as membrane association have been speculated to play a role in the possible mechanisms for substrate channeling between the active sites of these TFEs (Ishikawa et al, 2004;Sah-Teli et al, 2023. The reaction intermediates of the three TFE reactions are always negatively charged due to the phosphate groups of the CoA moiety and it has been proposed that positively charged residues on the surfaces of these proteins are used to guide the negatively charged intermediates between the three active sites.…”

Section: Introductionmentioning

confidence: 99%

“…In addition to the charged surfaces, also the formation of reaction chambers, due to the mode of assembly, has been proposed to play a role in substrate channeling. For HsTFE and anEcTFE membrane association is also hypothesized to be important in this respect (Sah-Teli et al, 2023;Xia et al, 2019).…”

Section: Introductionmentioning

confidence: 99%

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Crystallographic fragment binding studies of theMycobacterium tuberculosistrifunctional enzyme suggest binding pockets for the tails of the acyl-CoA substrates at its active sites and a potential substrate channeling path between them

Dalwani,

Metz,

Huschmann

et al. 2024

Preprint

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TheMycobacterium tuberculosistrifunctional enzyme (MtTFE) is an α2β2tetrameric enzyme in which the α-chain harbors the 2E-enoyl-CoA hydratase (ECH) and 3S-hydroxyacyl-CoA dehydrogenase (HAD) active sites, and the β-chain provides the 3-ketoacyl-CoA thiolase (KAT) active site. Linear, medium, and long chain 2E-enoyl-CoA molecules are the preferred substrates of MtTFE. Previous crystallographic binding and modelling studies have identified binding sites for the acyl-CoA substrates at the three active sites as well as the NAD+binding pocket at the HAD active site. These studies have also identified three additional CoA binding sites on the surface of MtTFE that are different from the active sites. It has been proposed that one of these additional sites could be of functional relevance for substrate channeling (by surface crawling) of reaction intermediates between the three active sites. Here, in a crystallographic fragment binding study with MtTFE crystals 226 fragments were screened, resulting in the structures of 17 MtTFE-fragment complexes. Analysis of the 143 fragment binding events shows that the ECH active site is the ′binding hotspot′ for the tested fragments, with 50 binding events. The mode of binding of the fragments bound at the active sites provides additional insight on how the long chain acyl moiety of the substrates can be accommodated at their proposed binding pockets. In addition, the 24 fragment binding events between the active sites identify potential transient binding sites of reaction intermediates relevant for possible channeling of substrates between these active sites. These results provide a basis for further studies to understand the functional relevance of these binding sites and to identify substrates for which channeling is crucial.

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Matching the β-oxidation gene repertoire with the wide diversity of fatty acids

Schiaffi,

Barras,

Bouveret

2024

Current Opinion in Microbiology

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Structural basis for different membrane-binding properties of E. coli anaerobic and human mitochondrial β-oxidation trifunctional enzymes

Cited by 4 publications

References 56 publications

DiffModeler: Large Macromolecular Structure Modeling in Low-Resolution Cryo-EM Maps Using Diffusion Model

DiffModeler: Large Macromolecular Structure Modeling in Low-Resolution Cryo-EM Maps Using Diffusion Model

Crystallographic fragment binding studies of theMycobacterium tuberculosistrifunctional enzyme suggest binding pockets for the tails of the acyl-CoA substrates at its active sites and a potential substrate channeling path between them

Matching the β-oxidation gene repertoire with the wide diversity of fatty acids

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