Overexpression, physicochemical characterization, and modeling of a hyperthermophilic Pyrococcus furiosus type 2 IPP isomerase

Raphaël Dutoit, Jérôme De Ruyck, Virginie Durisotti, Christianne Legrain, Eric Jacobs, Johan Wouters

Résultats de recherche: Contribution à un journal/une revueArticleRevue par des pairs

Résumé

In the first step of this study, type 2 isopentenyl diphosphate isomerase (IDI2) from Pyrococcus furiosus (pf-IDI2), a hyperthermophilic microorganism, was cloned and overexpressed in E. coli. After purification, hyperthermophilic behavior of this protein was approached by means of enzymatic assays and thermal denaturation studies. Compared with the mesophilic Streptococcus pneumoniae IDI2, which unfolds and looses activity above 50°C, pf-IDI2 is still folded and active at 80°C. Molecular modeling was applied, in a parallel step, to understand the molecular basis of thermal stability. Comparison of IDI2 from S. pneumoniae, T. thermophilus, and P. furiosus suggested that additional charged residues present in the hyperthermophilic enzyme might contribute to its higher thermal stability. This could increase the number of salt bridges between monomers of IDI2 in P. furiosus enzyme and, hence, decrease flexibility of loops or N-terminal segment, thereby enhancing its thermal stability.

langue originaleAnglais
Pages (de - à)1699-1707
Nombre de pages9
journalProteins: Structure, Function and Genetics
Volume71
Numéro de publication4
Les DOIs
Etat de la publicationPublié - 1 juin 2008

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