Psychrophilic protein modeling

Tubulin dimers of psychrophilic eukaryotes can polymerize into microtubules at 4°C, a temperature at which microtubules from mesophiles disassemble. This unique capability requires changes in the primary structure and/or in post-translational modifications of the tubulin subunits. To contribute to the understanding of mechanisms responsible for microtubule cold stability, here we present a computational structural analysis based on molecular dynamics (MD) and experimental data of three ?-tubulin isotypes, named EFBT2, EFBT3, and EFBT4, from the Antarctic protozoon Euplotes focardii that optimal temperature for growth and reproduction is 4°C. The unique characteristics of the primary and tertiary structures of psychrophilic ?-tubulin isotypes seem responsible for the formation of microtubules with distinct dynamic and functional properties. Folding assistance is a fundamental requirement for tubulin. Here, we report a pilot folding analysis of a divergent beta- tubulin isotype, named EFBT3, from the Antarctic psychrophilic ciliate Euplotes focardii. To attain its native monomeric structure, beta-tubulin needs the assistance of the eukaryotic class II chaperonin CCT and cofactor A (CofA). We demonstrated that the rare Cys281 of EFBT3 is critical for the folding reaction. Model predictions indicate that EFBT3 binds to CofA differently from yeast beta-tubulin, suggesting a diverse folding mechanism that may be correlated with microtubule cold adaptation. Superoxide dismutases (SODs) are ubiquitous enzymes, which catalyse the disproportion of superoxide to molecular oxygen and peroxide. This reaction preempts the oxidizing chain reaction, preventing several cell damage caused by reactive oxygen species. E. focardii has two Cu/Zn - SODs isoforms which display about 40% of sequence identity. In order to investigate the structural adaptation and the effect of ions at living low temperature and at mesophilic temperature, we performed a computational structural analysis based on molecular dynamics (MD) at 4°C and 27°C, and complexed or un-complexed with Cu and Zn ions, for comparing the ions effect on protein structure and the structure flexibility at different temperature.