CNR ExploRA

Articolo in rivista, 2002, ENG, 10.1074/jbc.M206761200

SDS-resistant Active and Thermostable Dimers Are Obtained from the Dissociation of Homotetrameric b-glycosidase From Hyperthermophilic Sulfolobus Solfataricus in SDS. Stabilizing Role of the A-C Intermonomeric Interface

Gentile F., Amodeo P., Febbraio F., Picaro F., Motta A. Formisano S., Nucci R.

F. Gentile, F. Picaro, S. Formisano: IEOS, CNR; P. Amodeo, A. Motta: ICB, CNR; F. Febbraio, R. Nucci: IBP, CNR.

beta-Glycosidases are fundamental, widely conserved enzymes. Those from hyperthermophiles exhibit unusual stabilities toward various perturbants. Previous work with homotetrameric beta-glycosidase from hyperthermophilic Sulfolobus solfataricus (M(r) 226,760) has shown that addition of 0.05- 0.1% SDS was associated with minimal secondary structure perturbations and increased activity. This work addresses the effects of SDS on beta-glycosidase quaternary structure. In 0.1-1% SDS, the enzyme was dimeric, as determined by Ferguson analysis of transverse-gradient polyacrylamide gels. The catalytic activity of the beta-glycosidase dimer in SDS was determined by in-gel assay. A minor decrease of thermal stability in SDS was observed after exposure to temperatures up to 80 degrees C for 1 h. An analysis of beta-glycosidase crystal structure showed different changes in solvent-accessible surface area on going from the tetramer to the two possible dimers (A-C and A-D). Energy minimization and molecular dynamics calculations showed that the A-C dimer, exhibiting the lowest exposed surface area, was more stabilized by a network of polar interactions. The charge distribution around the A-C interface was characterized by a local short range anisotropy, resulting in an unfavorable interaction with SDS. This paper provides a detailed description of an SDS-resistant inter-monomeric interface, which may help understand similar interfaces involved in important biological processes.

The Journal of biological chemistry (Print) 277 , pp. 44050–44060