Structural conservation in the glutathione binding in Sphingomonas sp. glutaredoxin Grx3 and variations for cold adaptation.

Glutaredoxin 3 (Grx3), a redox protein with a thioredoxin-fold structure, maintains structural integrity and glutathione (GSH) binding capabilities across varying habitat temperatures. The cis-Pro loop, essential for GSH binding, relies on the Arg-Asp salt bridge (α2-α3) and Gln-His hydrogen bond (ß3-ß4) for its conformation. In some psychrophilic Grx3 variants, Arg in α2 is replaced with Tyr, and His in ß4 is replaced with Phe. This study examines the roles of these bonds in Grx3's structure, function, and cold adaptation, using SpGrx3 from the Arctic bacterium Sphingomonas sp. Despite its cold habitat, SpGrx3 maintains the Arg51-Asp69 salt bridge and Gln56-His63 hydrogen bond. The R51Y substitution disrupts the α2-α3 salt bridge, while the H63F and H63Y substitutions hinder the salt bridge through cation-π interactions with Arg51, involving Phe63/Tyr63, thereby enhancing flexibility. Conversely, mutations that disrupt the hydrogen bond (Q56A, H63A, and H63F) reduce thermal stability. In the psychrophilic Grx3 configuration A48T/R51Y/H63F, a Thr48-Gln56 hydrogen bond stabilizes the cis-Pro loop, enhancing flexibility by disrupting both bonds. Furthermore, all mutants exhibit reduced α-helical content and catalytic efficiency. In summary, the highly conserved Arg51-Asp69 salt bridge and Gln56-His63 hydrogen bond are crucial for stabilizing the cis-Pro loop and catalytic activity in SpGrx3. His63 is favored as it avoids cation-π interactions with Arg51, unlike Phe63/Tyr63. Psychrophilic Grx3 variants have adapted to cold environments by reducing GSH binding and increasing structural flexibility. These findings deepen our understanding of the structural conservation in Grx3 for GSH binding and the critical alterations required for cold adaptation.

SATQPCR: Website for statistical analysis of real-time quantitative PCR data.

SATQPCR is a web tool providing statistical analysis of real-time quantitative PCR data including all MIQE rules (gene efficiency, selection of reference genes and normalization with them). Our application is a quick tool that provides to the biologist, graphs as well as statistical tables summarizing their results with the chosen methods (t-test or ANOVA with Tukey test). The application is available at http://satqpcr.sophia.inra.fr with a demo dataset. Source code can be found at https://framagit.org/. SUPPLEMENTARY INFORMATION: Tutorials at http://satqpcr.sophia.inra.fr/cgi/help.cgi.