bioRxiv Science⌕ Search
Home/Archive/bioRxiv

bioRxiv · 10.1101/2024.01.07.574536

Structural dynamics of the methyl-coenzyme M reductase active site are influenced by coenzyme F430 modifications

Poleto, M. D.·Allen, K. D.·Lemkul, J. A.
View original source ↗Export metadata ↓

Abstract

Methyl-coenzyme M reductase (MCR) is a central player in methane biogeochemistry, governing methanogenesis and the anaerobic oxidation of methane (AOM) in methanogens and anaerobic methanotrophs (ANME), respectively. The prosthetic group of MCR is coenzyme F430, a nickel-containing tetrapyrrole derivative. Additionally, a few modified versions of F430 have been discovered, including the 172-methylthio-F430 (mt-F430) that functions with ANME-1 MCR. This study employs molecular dynamics (MD) simulations to unravel the intricacies of the active-site dynamics of MCR from Methanosarcina acetivorans and ANME-1 when bound to the canonical F430 compared to 172-thioether coenzyme F430 variants and substrates for methane formation. Overall, our simulations indicate that each MCR active site is optimized for a given version of F430 and support the importance of the Gln to Val substitution in accommodating the 172 methylthio modification. Notably, modifications in the 172 position disrupt the canonical coordination among cofactors in M. acetivorans MCR, implicating structural perturbations, but evidence of active site reorganization to maintain substrate positions suggest that the modified F430s could be accommodated in a methanogenic MCR. We additionally report the first quantitative estimate of MCR intrinsic electric fields pivotal in driving methane formation. Our results suggest that the electric field aligned along the CH3-S-CoM thioether bond facilitates homolytic bond cleavage, coinciding with the proposed catalytic mechanism. Structural perturbations, however, weaken and misalign these electric fields, emphasizing the significance of the active site structure in maintaining their integrity. In conclusion, our results deepen the understanding of MCR active-site dynamics, the enzymes organizational role in intrinsic electric fields for catalysis, and the interplay between active site structure and electrostatics. This work not only advances our comprehension of MCR functionality but also provides a foundation for future investigations employing sophisticated models to capture the complex electronic properties of MCR active sites quantitatively.

Explore related subjects

biophysicsPreprint

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

Poleto, M. D., Allen, K. D., Lemkul, J. A.. 2024-01-08. Structural dynamics of the methyl-coenzyme M reductase active site are influenced by coenzyme F430 modifications. https://doi.org/10.1101/2024.01.07.574536

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

bioRxiv2026

Scaling of structural variability of ecDNA polymer condensates with copy number boosts and stabilises oncogene regulatory contacts

Extrachromosomal DNAs (ecDNAs) form highly heterogeneous condensates in cancer cells that drive oncogene overexpression, yet how structural variability coexists with stable gene regulation remains unclear. Here, we develop a minimal polymer physics model of MYC-harbouring COLO320-DM ecDNAs, where BRD4-like complexes bind and bridge cognate sites along ecDNA rings. Above a critical binder concentration, ecDNAs phase separate into condensates exhibiting diverse conformations because of their thermodynamic folding degeneracy. Despite this variability, condensates retain conserved interaction scaffolds that give rise to reproducible contact patterns, including in-trans associated domains (I-TADs), genomic regions enriched in intermolecular regulatory contacts between distinct ecDNAs. We find that condensate 3D architecture follows universal scaling relations with ecDNA copy number, n, remaining robust to model parameter changes. Regulatory contacts within I TADs increase linearly with n, yet they are one order of magnitude stronger than in size matched control regions outside I TADs, whereas their relative fluctuations are markedly suppressed as n increases. This scaling produces enhanced, low-noise regulatory environments for oncogenes embedded within I-TADs, such as PVT1-MYC fusions, whereas the canonical MYC copy, located outside, is less amplified as experimentally observed. Our findings reveal universal polymer physics principles underlying ecDNA condensate organization, offering a mechanistic basis for selective oncogene amplification and potential advantages in cancer progression.

biophysics↗
bioRxiv2026

High-resolution mapping of RNA structural maturation during Cas9 assembly with ABEL-FRET

The structural flexibility of RNA is essential for forming ribonucleoprotein (RNP) complexes, which regulate diverse biological processes. This intrinsic property permits RNA to act as a dynamic scaffold along the assembly pathway as it folds into a specific structure for initial recognition by protein and undergoes conformational rearrangements for functional maturation as a complex. Yet, RNA flexibility and RNP multicomponent assembly create significant obstacles for traditional structural methods. To overcome these challenges, we applied recently developed ABEL-FRET spectroscopy to measure tether-free single-molecule Forster resonance energy transfer (smFRET) over extended observation times. Furthermore, ABEL-FRET enables the unique ability for simultaneous measurements of ultrahigh resolution smFRET and hydrodynamic size of individual complexes, which offers distinct advantages for studying dynamic RNA molecules that undergo assembly via sequential binding events. Using ABEL-FRET, we explored how the guide RNA (gRNA) of CRISPR genome editing system folds and modulates its structural flexibility to carry out the roles required for each assembly state from its unbound apo form to the functional Cas9 RNP state for target DNA cleavage. Multi-perspective view of gRNA structure gained by probing its two primary functional domains enabled to capture dramatic changes in gRNA flexibility that are highly dependent on its specific structural domains as well as assembly states. Collectively, our work with ABEL-FRET highlights the intrinsic link between the structural flexibility of RNA and its functionality in RNP assembly.

biophysics↗
bioRxiv2026

De novo design of functional RNAs through higher-order interactions

Designing RNA sequences that reliably adopt functional three-dimensional structures remains a central challenge in RNA engineering because folding depends on cooperative interactions beyond canonical base pairing. Here we present DS3dRNA, an interaction-based framework for de novo RNA sequence design that combines a three-body statistical potential with physics-guided sequence sampling and supports design against multiple conformations. Across the evaluated benchmarks, DS3dRNA outperformed representative RNA inverse-design methods in native-sequence recovery and agreement between predicted and target structures. Energy-sequence-quality analyses further showed that lower design energies generally accompanied higher sequence recovery and macro-averaged F1 scores (MacroF1). Experimentally tested Mango II designs retained high-affinity fluorogenic activity, and five twister ribozyme designs yielded mean endpoint cleavage fractions of 37.7-50.6%, compared with 23.5% for the wild type. These results establish explicit higher-order interaction scoring as a complementary approach to emerging data-driven RNA design methods and provide a framework for designing functional RNAs from experimental or predicted structural ensembles.

biophysics↗