SDS-resistant active and thermostable dimers are obtained from the dissociation of homotetrameric beta-glycosidase from hyperthermophilic Sulfolobus solfataricus in SDS. Stabilizing role of the A-C intermonomeric interface.
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.
Gentile
Fabrizio
F
Istituto di Endocrinologia e Oncologia Sperimentale del CNR and Dipartimento di Biologia e Patologia Cellulare e Molecolare, Università Federico II, Via Pansini 5, 80131 Napoli, Italy.
Amodeo
Pietro
P
Febbraio
Ferdinando
F
Picaro
Francesco
F
Motta
Andrea
A
Formisano
Silvestro
S
Nucci
Roberto
R
D001665:Binding Sites; D019281:Dimerization; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D005959:Glucosidases / Q000737:chemistry* / Q000378:metabolism; D006358:Hot Temperature; D007700:Kinetics; D008958:Models, Molecular; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D012967:Sodium Dodecyl Sulfate / Q000494:pharmacology; D016944:Sulfolobus / Q000201:enzymology; D013696:Temperature
D012967:Sodium Dodecyl Sulfate; D005959:Glucosidases; C104851:Sulfolobus solfataricus beta-glycosidase