Biology subjects

Chenuel, T.

Publications and source records attributed to Chenuel, T..

4 recordsLinked to original sources

bioRxiv2026

Computational redesign of Chlamydomonas reinhardtii photosynthetic fructose-1,6-bisphosphatase from the Calvin-Benson-Bassham cycle improves both stability and catalytic capacities in vitro

Photosynthetic carbon fixation is catalyzed by 11 enzymes in the Calvin-Benson-Bassham cycle, a highly conserved metabolic pathway in the green lineage. Fructose-1,6-bisphosphatase (FBPase) contributes to the regeneration of the substrate of carboxylation. While non-photosynthetic paralogs of FBPases contribute to cytosolic neoglucogenesis in a well-documented manner, chloroplast FBPases specific regulations by phospho- and redox-posttranslational modifications, pH, cofactor binding, and homo-oligomeric exchange is only partially understood. In particular, FBPases from green microalgae are potentially target of a multiplicity of disulfide/dithiol exchanges that appear more complex than all their cytosolic and chloroplastic land-plant or cyanobacterial paralogs. In an attempt to decipher the molecular basis of chloroplast FBPase activities and precise redox regulations, we applied computational tools to extensively redesign its protein coding sequence, testing the simultaneous substitution of >13% of its amino acids as guided by distant homology. Resulting modified proteins in pure recombinant form were significantly more stable than the wild-type starting point, that was prone to denaturation or aggregation. In particular, redesigned FBPase favored the catalytically competent homo-tetrameric state. Thermal denaturation assays revealed that the redesign of FBPase sequence starkly increased in thermostability, in good correlation with an augmented enzymatic activity over long periods of time. By comparison with its wild-type counterpart, computationally redesigned FBPase allowed an extensive structure-function study that uncovered the multiplicity of redox regulations of microalgal paralog, their combined contribution to the assembly of the catalytically competent homo-tetrameric state, and will further facilitate molecular engineering of photosynthesis.

biochemistry↗

bioRxiv2026

Structural basis of HSP90C, a highly active chloroplastic HSP90 chaperone from A. thaliana

Chloroplasts are the main energy organelles in plants, primary through photosynthesis. Thereby, they are responsible for CO2 fixation and dioxygen production, which are essential for living species on Earth. To ensure these processes, numerous proteins encoded from the nuclear DNA need to be imported inside the chloroplast, and eventually to the thylakoids. Whereas the translocation systems from both chloroplastic and thylakoids membranes have been studied in recent years, the stromal route between these two membranes is largely unknown. Notably, the chloroplastic HSP90 (HSP90C) is likely to play an important role in this process, but its structure and molecular mechanisms remain to be unveiled. In this study, we used a combination of structural and biophysical approaches to elucidate the features of Arabidopsis thalianas HSP90C. Principally, we found that HSP90C has a remarkably high ATPase activity among the HSP90 family proteins. Further investigation allowed us to pinpoint atypical mechanisms responsible for this high activity. First, the N-terminal cap is involved in a disulfide bond that accelerates the ATPase activity of HSP90C. Second, its C-terminal domain features an extension that is mandatory for its dimerization. Third, our crystal structures reveal a wide opening of the HSP90Cs dimer with reduced intermonomeric interfaces. Lastly, we identified a helical switch which is required for HSP90Cs high activity. Three of these four features are due to sequence signatures of HSP90C, which we found to be shared by most of green plants representatives. Our study provides first insights of HSP90Cs non-canonical mechanisms, which will help in the understanding of processes related to protein import in the chloroplast.

biochemistry↗

bioRxiv2025

R2TP-like Quaternary Chaperones: a comprehensive overview to understand the dynamic R2SP complex

The human R2SP complex belongs to the R2TP-like quaternary chaperone family and consists of RUVBL1, RUVBL2, SPAG1 and PIH1D2. R2SP is crucial for the correct assembly of motile cilia (SPAG1 null mutations cause Primary Ciliary Dyskinesia) and the organization of the synaptic zone. RUVBL1/2 ATPases are the powerhouse of this molecular machinery, while SPAG1 and PIH1D2 would be adaptors that interact with specific clients to promote their quaternary assembly. Despite these functional data, little is known about the structure of R2SP and the precise mode of action of these R2TP-like complexes. We have combined biochemical and structural approaches (NMR, structural mass spectrometry and cryo-EM) to investigate the 3D organization of the human R2SP complex, its mode of assembly and ATPase activity. Our study reveals a three-dimensional structure similar to that of the canonical R2TP complex, but also highlights differences in the mode of action of its RUVBL1/2 ATPase core as well as the binding of its adaptors SPAG1 and PIH1D2.

biophysics↗

bioRxiv2023

Identification of the Intracellular Protein Targets of a Bio-active Clickable Half-Sandwich Iridium Complex by Chemical Proteomics

Identification of intracellular targets of anticancer drug candidates provides key information on their mechanism of action. Exploiting the ability of the anticancer (C^N)-chelated half-sandwich iridium(III) complexes to covalently bind proteins, click chemistry with a bioorthogonal azido probe was used to localize a phenyloxazoline-chelated iridium complex within cells and profile its interactome at the proteome-wide scale. Proteins involved in protein folding and actin cytoskeleton regulation were identified as high affinity targets. Upon iridium complex treatment, HSP90 folding activity was inhibited in vitro and major cytoskeleton disorganization was observed. We used a wide array of imaging and biochemical methods to validate selected targets and obtain a multiscale overview of the effects of this complex on live human cells. We demonstrate that it behaves as a dual agent, inducing both electrophilic and oxidative stresses in cells that account for its cytotoxicity.

biochemistry↗