title: Small heat shock protein Hsp27 prevents heat-induced aggregation of F-actin by forming soluble complexes with denatured actin.
abstract: Previously, we have shown that the small heat shock protein with apparent molecular mass 27 kDa (Hsp27) does not affect the thermal unfolding of F-actin, but effectively prevents aggregation of thermally denatured F-actin [Pivovarova AV, Mikhailova VV, Chernik IS, Chebotareva NA, Levitsky DI & Gusev NB (2005) Biochem Biophys Res Commun331, 1548-1553], and supposed that Hsp27 prevents heat-induced aggregation of F-actin by forming soluble complexes with denatured actin. In the present work, we applied dynamic light scattering, analytical ultracentrifugation and size exclusion chromatography to examine the properties of complexes formed by denatured actin with a recombinant human Hsp27 mutant (Hsp27-3D) mimicking the naturally occurring phosphorylation of this protein at Ser15, Ser78, and Ser82. Our results show that formation of these complexes occurs upon heating and accompanies the F-actin thermal denaturation. All the methods show that the size of actin-Hsp27-3D complexes decreases with increasing Hsp27-3D concentration in the incubation mixture and that saturation occurs at approximately equimolar concentrations of Hsp27-3D and actin. Under these conditions, the complexes exhibit a hydrodynamic radius of approximately 16 nm, a sedimentation coefficient of 17-20 S, and a molecular mass of about 2 MDa. It is supposed that Hsp27-3D binds to denatured actin monomers or short oligomers dissociated from actin filaments upon heating and protects them from aggregation by forming relatively small and highly soluble complexes. This mechanism might explain how small heat shock proteins prevent aggregation of denatured actin and by this means protect the cytoskeleton and the whole cell from damage caused by accumulation of large insoluble aggregates under heat shock conditions.
authors.affiliation: A. N. Bach Institute of Biochemistry, Russian Academy of Sciences, Moscow, Russia.
authors.forename: Anastasia V
authors.initials: AV
authors.lastname: Pivovarova
authors.forename: Natalia A
authors.initials: NA
authors.lastname: Chebotareva
authors.forename: Ivan S
authors.initials: IS
authors.lastname: Chernik
authors.forename: Nikolai B
authors.initials: NB
authors.lastname: Gusev
authors.forename: Dmitrii I
authors.initials: DI
authors.lastname: Levitsky
mesh_terms: D000199:Actins / Q000737:chemistry / Q000378:metabolism*; D000818:Animals; D002850:Chromatography, Gel; D006360:Heat-Shock Proteins / Q000737:chemistry / Q000378:metabolism*; D011489:Protein Denaturation; D011817:Rabbits; D014461:Ultracentrifugation
chemical_list: D000199:Actins; D006360:Heat-Shock Proteins